MXPA99010851A - Use of recombinant human uteroglobin in treatment of inflammatory and fibroticconditions - Google Patents

Abstract

Methods for treatment of inflammatory and fibrotic conditions in vivo using UG is disclosed. Methods for treating or preventing inflammatory or fibrotic conditions characterized by a deficiency of endogenous functional UG are also disclosed. Compositions containing UG, optionally containing lung surfactant, and assay procedures for detection of UG-fibronectin complexes, are also provided.

Description

USE OF RECOMBINANT HUMAN UTEROGLOBIN IN THE TREATMENT OF INFLAMMATORY AND FIBRAL TREATMENTS REFERENCE TO RELATED APPLICATIONS The present application for a continuation in part of the United States application serial No. 08 / 864,357, the description of which is incorporated herein by reference.

FIELD OF THE INVENTION The invention relates, in general, to the treatment of inflammatory and fibrotic conditions using native human uteroglobin (hUG) or recombinant human uteroglobin (rhUG). Physiological functions and novel therapies for UG (hUG or rhUG) have been identified. Specifically, the invention relates to the treatment of inflammatory and fibrotic conditions by the administration of hUG or rhUG to inhibit PLA2 and / or to prevent the deposition of fibronectin. The invention also provides a method of treating neonatal respiratory distress syndrome (SPR) and bronchopulmonary dysplasia (BPD), a critical clinical state of the lung, and glomerular nephropathy, a kidney disease, both characterized by inflammatory and fibrotic conditions.

The documents mentioned in this application relate to the state of the art to which this invention pertains, each of which is incorporated herein by reference.

BACKGROUND OF THE INVENTION Inflammatory and fibrotic diseases The search for improved therapeutic agents for the treatment of inflammatory and fibrotic diseases has recently received great attention. The neonatal SPR, a disease of surfactant deficiency in the lung is a condition of particular interest in that it is one of the leading cases of mortality in preterm infants. Although the introduction of surfactant therapy drastically improves the survival of patients with SPR, the development of chronic inflammatory and fibrotic disease in a significant percentage of this patient population is a major problem. In the same way, the glomerular nephropathy with fibronectin deposit, hereditary, gives rise to a terminal renal failure when the kidney of the patients is blocked and the blood is no longer filtered. Nephropathy is characterized by deposits of fibronectin and fibrosis of the kidneys that returns to the non-functional organ and, finally, unable to support life.

PLA2 (phospholipase A2), a class of endogenous enzymes that hydrolyze the ester bond in the Sn2 position of glycerophospholipids, is one of the many proteins involved in inflammatory and fibrotic conditions. It is also responsible for the hydrolysis of surfactant phospholipids in the lungs. Uteroglobin (also known as CC10, CC16, CC17, urine protein-1, Pl, progesterone binding protein, PCB binding protein, Clara cell secretory protein (CCSP), blastokinin, retinol binding protein, protein binding to phospholipids and alpha2-microglobulin) inhibits the activity of PLA2 in vi tro. Uteroglobin is a small, globular homodimeric protein. It has a molecular weight of 15.8 kDa, but migrates in electrophoretic gels to a size corresponding to 10 kDa. Human uteroglobin is abundant in the adult human lung and contains up to about 7% of the total soluble protein. However, its expression is not fully activated in the developing human fetus until the last stage of gestation. As a result, the extracellular fluids of the lung of newborns before the term contains much less UG than of adults. The UG is also expressed by the pancreas. PLA2 plays crucial roles in the inflammatory response because they release arachidonic acid (AA) from the cellular reservoirs of phospholipids. AA is metabolized to a number of potent inflammatory mediators in a process called arachidonic acid cascade. Various acute and chronic clinical conditions have been characterized by the elevated activity of PLA2 in serum or local (see Table 1 below).

Table 1. Clinical conditions associated with the activity of PLA2 respiratory disease syndrome in adults At present there are no effective PLA2 inhibitors, available for clinical use. To date, only some PLA2 inhibitors have progressed in clinical trials, but none have qualified for commercial use. Fibronectin (Fn) is a 200 kDa glycoprotein that It exists in some different forms and is secreted by different tissues. Fn is an essential protein and directed to the breakdown of the Fn gene in mice showed that it has a main function in embryogenesis. Fn also plays a key role in inflammation, cell adhesion, tissue repair and fibrosis, and is deposited at the site of the injury. Plasma fibronectin (pFn) is secreted by the liver and circulates in the plasma. In the lung, cellular Fn (cFn) is secreted with inflammation and injury. Both types of Fn are chemotactic factors for inflammatory cells and fibroblasts. Large amounts of inflammatory cells and fibroblasts infiltrate the lung during inflammatory episodes, which can lead to pulmonary fibrosis and ultimately death. High levels of Fn have been detected in clinical conditions in humans such as SPR and neonatal LBP of the lung, and glomerular nephropathy of the kidney.

The function of the UG The amino acid analysis of the purified human UG reveals that it is structurally similar, but not identical, to other "UG-like" proteins, for example, the rabbit UG 39 of 70 amino acids are identical between UG of human and rabbit (see Figure 1).

"UG-like" including human UG / CC10, rat CC10, mouse CC10, and rabbit UG exhibit species-specific and tissue-specific antigenic differences, as well as differences in tissue distribution and in vitro biochemical activities. UG-like proteins have been described in many different contexts with respect to tissues and species of origin, including rat lung, human urine, sputum, blood components, rabbit uterus, rat and human prostate, and human lung . At present there are no known physiological functions for these proteins. Despite the years of study, the biological functions of these proteins in vivo remain unclear. The absence of structural identity between the UG-like proteins makes it impossible to predict whether a protein will possess therapeutic function in vivo in humans based on in vitro activity or another presented by a structurally related protein. For example, human uteroglobin binds less than 5% of the amount of progesterone compared to the rabbit UG in the same assay. The human UG has a lower isoelectric point (4.6) compared to the rabbit UG (5.4). Stripp et al (1996) have reported studies on a mouse treated with uteroglobin generated to eliminate expression of uteroglobin. The mouse had Clara cells that they present singular intracellular structures instead of uteroglobin secretion granules, but no other phenotype exists. This observation is highly significant because lung function accompanied by lung inflammation and fibrosis was expected. In addition, this mouse faint showed no evidence of renal, pancreatic or reproductive abnormality indicating that the uteroglobin protein had no significant function in controlling inflammation or fibrosis in vivo.

OBJECTS OF THE INVENTION Therefore, it is an object of the present invention to provide a pharmaceutical composition that includes an effective amount that inhibits PLA2 from recombinant human uteroglobin (rhUG) or a fragment or derivative thereof and a pharmaceutically acceptable carrier or diluent. Another object of the invention is to provide a pharmaceutical composition which includes an amount effective to bind recombinant human uteroglobin fibronectin or a fragment or derivative thereof and a pharmaceutically acceptable carrier or diluent. A further objective of the present invention is to provide a pharmaceutical composition that includes an amount effective to inhibit PLA2 or bind fibronectin to an active agent for the treatment of an indication objective. Furthermore, an object of the invention is to provide a method for inhibiting PLA2 enzymes in vivo in a mammal in need of this treatment, wherein the method includes administering to a mammal an PLA2 inhibitory effective amount of native or recombinant human uteroglobin or a fragment or derivative thereof. Still further, it is an object of the present invention to provide a method for the treatment or prevention of an inflammatory condition in a patient in need of such treatment, wherein the method includes the administration of an anti-inflammatory effective amount of native or recombinant UG or a fragment or derivative thereof.

It is an object of the present invention to provide a method for the treatment or prevention of a fibrotic condition in a patient in need of such treatment, wherein the method includes administering an effective amount to bind fibronectin from native or recombinant UG or a fragment or derivative thereof. A further object of the present invention is to provide a method for the treatment or prevention of an inflammatory or fibrotic condition characterized by a deficiency of endogenous UG, wherein the method includes the administration of a compensatory amount of the native UG or recogminant or a fragment or derived from it.

Another object of the present invention is to provide a cosmetic composition containing an effective inhydride amount of PLA2 or to bind native or recombinant UG fibronectin, or a fragment or derivative thereof. In addition, it is an object of the present invention to provide a blood supplement containing an effective PLA2 inhibiting amount or to bind native or recombinant UG fibronectin or a fragment or derivative thereof. Finally, an object of the present invention is to provide an assay for the quantification of uteroglobin-fibronectin complexes in a chemical sample.

SUMMARY OF THE INVENTION Uteroglobin has now been found to play a central physiological role in the inhibition of PLA2 in the prevention of fibronectin deposition and fibrosis in vivo. A combination of the experiments performed on a new strain of transgenic mice "knocked out or passed out" with uteroglobin, and a model of neonatal respiratory distress syndrome (SPR) in mono that includes inflammation and pulmonary fibrosis demonstrates these effects. The mouse fainted with uteroglobin of the present invention (hereinafter the "KO UG mice / mouse") exhibits lethal glomerular nephropathy and renal parenchymal fibrosis, as early and late onset diseases, respectively. The administration of exogenous Fn to normal mice causes the deposition of Fn in the kidneys, but the administration of equimolar amounts of Fn and rhUG do not present this deposit. The reduction of PLA2 activity in vivo has been demonstrated in the presence of rhUG. In a first experiment, the mouse KO UG phenotype showed that the activity of PLA2 in serum rises significantly in the absence of UG, compared to the activity of PLA2 in litters that present a functional UG gene. In a second experiment, the administration of rhUG to pre-term monkeys suffering from SPR showed the inhibition of PLA2 activity in the extracellular fluids of the lungs. Other experiments show that PLA2 in vitro can degrade the artificial surfactant (usually Survanta) used in the treatment of SPR and that UG can inhibit this degradation. These experiments demonstrate that UG mediates the inhibition of PLA2 and the deposition of Fn in vivo after intratracheal or intravenous administration. Experiments with mice fainted with uteroglobin show that rhUG can be used to treat conditions in which uteroglobin is deficient or the protein itself carries a mutation of loss of function. Now it has been discovered that the rhUG can be used to treat or prevent inflammatory or fibrotic conditions in which functional endogenous uteroglobin is deficient in the circulation or at the site of inflammation or fibrosis. In certain pulmonary inflammatory or fibrotic conditions, reductions in serum hUG levels and / or bronchoalveolar lavage fluids have been found, including preterm infants at risk of developing neonatal BPD. It has been found that UG can be used to supplement defective or defective endogenous uteroglobin to prevent or treat these inflammatory and fibrotic conditions. According to one aspect, the invention provides a method of treating an inflammatory condition in vivo, which comprises administering to a patient in need of this treatment an anti-inflammatory effective amount of UG. According to another aspect, the invention provides a method for the inhibition of soluble PLA2 enzymes in vivo, which consists of administering to a patient in need of this treatment an amount of UG effective to inhibit PLA2. According to yet another aspect, the invention provides a method for the treatment or prevention of a fibrotic condition, which consists of administering to a patient in need of this treatment, an amount of UG effective for binding to fibronectin. Another aspect of the invention provides a method of treating or preventing fibrillogenesis by adding an amount of UG for binding to fibronectin. According to another aspect, the invention provides a method for the treatment or prevention of an inflammatory or fibrotic condition characterized by a deficiency of functional, endogenous UG, which consists in administering to a patient in need of such treatment a compensating amount of UG. The invention also provides pharmaceutical compositions containing an effective amount of rhUG in association with a pharmaceutically acceptable carrier or diluent. The compositions may take the form of injectable and liquid or semi-aerosol solutions for intratracheal administration. According to another aspect, the invention provides pharmaceutical compositions containing UG and a lung surfactant (for example Survanta (extract from bovine lung from Abbot Labs) and Exosurf (a lung surfactant obtained by chemical synthesis of Glaxo-Wellcome)) in association with a pharmaceutically acceptable carrier or diluent. The invention also includes the compositions Pharmaceuticals containing an effective amount to inhibit PLA2 or for the binding to fibronectin of rhUG, an active agent for the treatment of an objective indication and a carrier. The effective amount of rhUG for the inhibition of PLA2 with binding to fibronectin reduces inflammation and by this means ensures that an effective amount of the active agent reaches the treatment site. In another aspect, the invention provides an assay for quantifying uteroglobin-fibronectin complexes in a clinical sample, wherein the clinical specimen suspected to contain a uteroglobin-fibronectin complex is contacted with an agent that captures the antigen, for example, a monospecific rabbit polyclonal antibody is immobilized on an insoluble support; an agent for the detection of the antigen is added to the sample, for example, an antibody specific for fibronectin; and the presence of any complex bound to the support is detected using, for example, a secondary antibody, for example anti-IgG antibody conjugated to an enzyme such as horseradish peroxidase using a normal enzymatic reaction, wherein the substrate of the enzyme is converts it into a chromogenic or fluorogenic compound that is quantified using standard spectrophotometric or fluorometric apparatus. The invention also provides a composition Cosmetic and a blood supplement that includes an effective amount of human uteroglobin to inhibit PLA2 and / or to bind to fibronectin and a pharmaceutically acceptable carrier or diluent.

BRIEF DESCRIPTION OF THE DRAWINGS The invention will now be described in greater detail with reference to the accompanying drawings, in which: FIGURE 1 shows an alignment of UG-like proteins; FIGURE 2A shows the proposed target construction of the mouse fainted with transgenic UG; the restriction sites are B-BamIII, E = EcoRI, H = HindIII; FIGURES 2B-2D show the verification of the genetic construction in the progeny of transgenic embryos by PCR and Southern blot analysis; (B) Southern blot analysis of the directed Rl ES cell clones, where Wt = wild-type; (C) representative PCR analysis of genomic DNA from tail biopsies of the offspring; the genotypes and their corresponding PCR products are as follows: UG + / +, 304 bp; UG + ", 304 and 667 bp; UG_ /", 667 bp; (D) Southern blot of mouse tail genomic DNA; FIGURE 2E shows confirmation of the absence of UG-mRNA in the lung tissues of UG ~ _ mice by RT-PCR analysis; analysis of the RT / PCR of the total RNA extracted from the bait lung tissues of genotypes UG + / +, UG and UG; a product of the RT / PCR of 273 bp was detectable in the lungs of UG + / + and UG + / "but absent from the UG-" mice; FIGURE 2F shows the confirmation of the absence of the UG protein in the lungs of UG_ ~ mice by Western analysis; proteins (30 micrograms each) from Lung Were subjected to electrophoresis using gradient 4/20% SDS-PAGE under non-reducing conditions and subjected to immunoassay using rabbit anti-mouse UG; FIGURE 2G shows confirmation of the absence of UG in sections of lung tissue from UG_ "mice using immunohistochemical methods in bronchiolar epithelial cells; dark staining over bronchiolar epithelial cells from UG + + mice (upper panel) indicates immunoreactivity of the UG, observe the absence of immunoreactivity in the lungs of UG "'" mice (lower panel) FIGURES 3A-3J compare the histopathological analyzes of kidney sections from normal mice compared to UG _, _, showing abnormal parenchymal fibrosis and glomerular Fn deposition only in faint mice, H & E staining of kidney sections from a UG + / + (A) and its bait partner UG_ / ~ (B); (C) kidney section of a 10-month mouse with severe parenchymal fibrosis (D) a region of the same mouse kidney in (C) showing renal tubular hyperplasia (40X amplification, g = glomeruli, f = fibrosis, t = tubule); (E) transmission electron microscopy of the glomerular deposit of a mouse UG "_ with severe renal disease (6000X amplification); (F) the start at (E) is amplified 60,000X, which shows the fibrillated structures, long indicative of collagen (col) and some short diffusions consistent with Fn fibrils; (G) Fn immunofluorescence of a kidney section from mouse UG + + using murine Fn antibody; (H) Fn immunofluorescence of a kidney section from a UG- / ~ mouse with severe renal disease, Mason trichrome staining of the kidney sections with UG + / + (I) and UG ~ _ (J) mice, the blue stain on the glomeruli of the mouse kidney section UG_ "It is collagen (approximately 40X amplification). FIGURE 4A shows the presence of Fn aggregates only in the kidneys of the UG_ / "mice, the immunoprecipitation and the Western blot analysis of the Fn from plasma, kidney and liver of UG ++ + and UG ~ / _ mice; Multimeric Fn (arrow in bold) was detected only in the kidney kidney Used UG_ ~ FIGURES 4B and 4C show the formation of the UG / Fn complexes in vitro, (B) equimolar concentrations of UG and Fn were incubated, immunoprecipitated and detected by Western blot with Fn or UG antibody; the immunoprecipitates contain Fn (lane 2, upper panel) and UG (lane 2, lower panel); strips 1 of the panels represent the standards of Fn and UG; (C) Equimolar concentrations of 125 I-UG and Fn were incubated at 4 ° C for one hour and the resulting complex was subjected to electrophoresis in 6% non-reducing, non-denaturing polyacrylamide gels; Strip 1, Coomassie blue, stained a heteron of Fn-UG; Lane 2, your autoradiogram. FIGURE 4D shows the presence of the UG-Fn complexes in the plasma of normal mice but not UG ~ / -; immunoprecipitation of plasma from UG + / + and UG ~ _ mice with Fn-antibody and Western blot analysis with Fn and UG antibodies; Fn (upper panel); UG (lower panel); std = standards for UG and Fn. FIGURE 4E shows the dose-dependent inhibition of autoaggregation of Fn by UG in vitro; affinity crosslinking of 125 I-Fn with unlabeled Fn in the absence (lane 2) and presence of different amounts of UG (lanes 3-5); the very high molecular weight intensity, radioactive Fn band (lane 2) formed in the absence of UG is reduced in a dose-dependent manner; lane 1, 125 I-Fn with unlabeled Fn in the absence of UG and DSS; clear arrowhead-multimeric Fn; thin lower arrow = 220 kDa of Fn. FIGURE 4F shows the inhibition of the formation of the Fn-collagen complex by UG; affinity crosslinking of 125I-collagen with unlabeled Fn in the absence of (band 3) and presence (band 4) of UG; lane 1, collagen I stained with Coomassie blue; alphai-alphai chain of collagen I and alpha2-alpha2 chain of collagen I; lane 2, 125I-collagen I and Fn not labeled in the absence of UG and DSS. FIGURES 5A-5F show the immunohistochemical analysis of the deposit of Fn in the kidneys of normal mice and UG "_ only in the absence of UG; (A) kidney section of a wild-type mouse that received a mixture of equimolar concentrations of Fn and UG intravenously; (B) UG + + mouse that received the same dose of Fn as in (A) but without UG; (C) UG mouse _ / _ apparently healthy receiving a mixture of Fn and UG; (D) mouse UG_ "receiving Fn alone (the same dose as in (C)), but without UG; (E) Fn-fibrillogenesis by cultured cells grown in medium supplemented with hFn alone; (F) a cell culture identical to that it is shown in (E) that it was fed with medium containing a mixture of equimolar concentrations of soluble hFn and UG (40X amplification, g = glomerulus) FIGURES 6A-6B show the format of a diagnostic test to detect complexes UG-Fn in clinical samples FIGURE 7 shows the passage of UG dimer through a dialysis membrane MWCO of 8.0 kDa but not by a MWCO dialysis membrane of 3.5 kDa.

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS rhUG The rhUG of the invention has practically the same amino acid sequence as the native human UG protein. An amino acid sequence having "virtually the same" amino acid sequence as that of the native human protein includes rhUG having at least 75% identity to the native human protein. In a preferred embodiment, the rhUG has at least 85% identity, and in a more preferred embodiment, the rhUG has at least 98% identity to the native UG. Also included in the method of the present invention is the use of UG fragments or derivatives. A "fragment" of UG refers to a portion of the amino acid sequence of the native hUG having 6 or more contiguous amino acids of the native protein sequence. The term "derivative" refers to peptide analogues of UG, which includes one or more amino acid substitutions and / or the addition of one or more chemical moieties, for example, acylating agents or sulfonating agents, with the proviso that the derivative retain the biological activity of the source molecule. In addition, the UG used in the method of this invention is substantially pure. The term "substantially pure" refers to the UG having a purity of about 75% to about 100%. In a preferred embodiment, the UG has a purity of about 90% to about 100%, and in the most preferred embodiment, the UG has a purity of at least 95%.

Clinical Uses of UG The invention provides, in another aspect, a method of treatment or prevention of an inflammatory or fibrotic condition which consists of administering to an animal, which can be animal or human, an effective amount of UG. The following non-limiting list of conditions are representative examples of those associated with UG deficiencies, excessive PLA2 activity and fibronectin deposition.

Table . Clinical uses of recombinant uteroglobin (grouped by the properties of UG) The common relationships between UG deficiency, PLA2 activity and aggregation and deposition of fibronectin in inflammatory / fibrotic conditions in humans are summarized below.

Neonatal bronchopulmonary dysplasia (neonatal BPD) Neonatal BPD is characterized by severe inflammation and irreversible fibrosis of lung tissue in newborns, usually as a result of respiratory distress syndrome (SPR). However, this condition can also be caused by meconium aspiration syndrome or infection. In this condition a deficiency of hUG has been implicated because the synthesis of pulmonary hUG can be co-regulated with the surfactant, which begins in the last stage of pregnancy. Thus, severely premature neonates may lack UG as well as surfactant. HUG deficiency may cause increased activity of PLA2 and fibrosis related to Fn, which are associated with inflammation and fibrosis observed in neonatal BPD. Some newborns do not respond to synthetic surfactant, which may be due to excess activity of PLA2. In this way, the UG can be used to treat neonatal BPD. Direct administration by endotracheal or systemic routes is the preferred route of administration.

Multiple Organ Failure (BMD) Excessive activity of PLA2 has been implicated in BMD due to bacterial sepsis or trauma. This condition is characterized by a systemic inflammatory response, rapid complication, massive damage to tissue and loss of organ function in the lungs, kidney, pancreas, intestines and vasculature. Recent evidence indicates that BMD is activated as the activity of soluble, systemic phospholipase A2 increases, its direct lysis of tissue cell membranes and hydrolysis of essential phospholipids, such as lung surfactants. Attempts to inhibit PLA2 directly in clinical settings have been unsatisfactory. In BMD, the amount of endogenous UG is insufficient to counteract the overactivation of PLA2. UG administered exogenously can be used to fight BMD. Insufficient remote organ (DOR) includes damage to organs other than the organ primarily affected by trauma or infection. Often the inadequacy of the remote organ includes more than a remote organ, resulting in multiple organ failure. For example, pancreatitis is an inflammation of the pancreas in response to alcohol ingestion, infection or trauma, which can lead to respiratory distress syndrome in adults (ARDS), acute renal failure (AKI), and systemic shock. The episode of inflammatory bowel disease or peritonitis can give rise to DOR / BMD. The DOR / BMD is associated with elevated levels of activated, circulating PLA2. The systemic application of hUG can avoid DOR / BMD. The immediate injection of UG in patients with DOR / BMD could reduce the severity or eliminate organ failure mediated by PLA2 and shock.

Pancreatitis All forms of pancreatitis include elevated activity of soluble PLA2, type I, systemic and local. Pancreatitis often leads to pulmonary insufficiency or ARDS, characterized by elevated activity of soluble PLA2 in the lungs. Therefore, as an inhibitor of type I PLA2, soluble, in vivo, UG is an excellent candidate for the treatment of two forms of acute pancreatitis, and as a preventive measure of pulmonary insufficiency in all acute forms of pancreatitis. The preferred route of administration is intravenously.

Inflammed bowel disease Inflammatory bowel disease (ElI), which includes ulcerative colitis, directiculitis and Crohn's disease, is characterized by high local production and activity of type II soluble PLA2. The activity of circulating soluble PLA2 may also be elevated in IBD. IBD causes pulmonary insufficiency or ARDS in severe cases, as a result of the elevated activity of PLA2 (similar to pancreatitis). The rationale for l-a-application of exogenous UG in ElI is identical to that of pancreatitis: deregulate the inflammatory response by inhibiting PLA2, aggregation and / or deposition of Fn and prevent evolution in a remote organ (lungs and kidneys). The preferred route of administration is intravenously in hospitalized patients.

Bacterial pneumonia BAL fluids from patients who had survived bacterial pneumonia showed higher levels of 2-3X UG who died. Bacterial infection of the lungs can overactivate soluble, endogenous PLA2. UG can be administered to inhibit or control this effect. The preferred route of administration is the intratracheal route if the patient is intubated or intravenous if it is not.

Complications of dialysis The main complication of dialysis is thrombosis, that is, the spontaneous formation of blood clots. These often block the entry of vascular access, deteriorating treatment, as well as causing ischemic episodes, sometimes threatening the patient's life. A second problem with hemodialysis patients is the inflammation and / or fibrosis of the proximal vein that returns the dialyzed blood to the main circulation of the patient. Proximal vein fibrosis is usually detected as an increase in resistance, or pressure, against the return of dialyzed blood. A third problem is fibrosis and closure of the vascular access site, or fistula. A fourth problem is accelerated atherosclerosis and a fifth is the loss of residual renal function, most likely due to the deposition of Fn. The possibility that the endogenous UG is dialyzed during the procedure provides an explanation for these problems. The selective elimination of the endogenous UG leaves the circulating Fn free to aggregate, forming the foci for the formation of blood clots or to be deposited on the erythrocytes, priming them for a coagulation response by adhesion between these or the vascular lumen. Transglutaminases (TG) are enzymes responsible for the formation of macromolecular reticles found in basal membranes, skin and blood clots. In the absence of free UG competing as a substrate for activated TGs, Fn and other components of blood clots are cross-linked.

The inflammation and fibrosis of the proximal vein and the vascular access site, as well as the accelerated atherosclerosis can be explained by the deposit of Fn in the vascular lumen. The deposit of fibronectin in the vascular endothelium favors the adhesion of platelets and leukocytes, which can be aggravated in the absence of PLA2 inhibition. The vascular deposits of Fn can also favor the local deposits of fat, cholesterol and proteins found in the atherosclerotic plaque. It is known that fibronexion is a major component of atherosclerotic plaque, as well as renal glomerular deposits associated with nephropathy and loss of primary and residual renal function. Therefore, UG administration can reduce or eliminate these problems by reducing inflammation and fibronectin deposition. The preferred route of administration of UG would be intravenous infusion before, during or after dialysis. Otherwise, the loss of endogenous UG can be prevented by adding UG to the dialysis buffer or by pre-debonding the dialysis membrane with UG or both.

Organ transplants The term "organ" refers, for example, to solid organs such as kidney, liver and heart, as well as bone marrow, cornea and skin.

There are two types of organ transplant rejection: acute and chronic. Acute rejection is an inflammatory process that involves the activity and infiltration of PLA2 by inflammatory cells that often destroy the graft. Chronic rejection involves fibrosis of the Fn-mediated graft, including atherosclerosis confined to the graft. Thus, the administration of UG can be used to treat or prevent acute or chronic rejection of the graft. The preferred route of administration is by injection. Another aspect of organ transplantation is ischemia of the organ before it is separated from the donor, during transport and in the recipient, which contributes to acute rejection. It is known that ischemia gives rise to an elevated activity of PLA2 and tissue necrosis. Therefore, the UG can be used to avoid these ischemia. Hence, the UG can be used to prevent ischemia. The preferred form of UG is as a liquid for perfusion or storage buffer in which the ex vivo organ is conserved.

Prevention of type I diabetes Type I diabetes arises from the destruction of pancreatic tissue by an autoimmune response. The pancreas normally secretes soluble PLA2 and hUG into the circulation.

Necrotic lesions have been reported in the pancreas of the knocked out mice (KO) with uteroglobin of the present invention (herein referred to as the "KO UG mouse") In the absence of uteroglobin, the KO UG mouse exhibits similar pancreatic tissue destruction as it could activate an autoimmune response. Thus, UG can be used to prevent or impede the slow progress of type I diabetes. The preferred route of administration is by injection.

Prevention and treatment of nephropathy Fn renal and fibrosis deposits in KO mice UG are similar to the deposits of Fn and fibrosis in human nephropathies. Thus, the administration of UG can prevent or slow down the progress of nephropathy in patients at risk, such as type II diabetes.

Prevention and treatment of ocular inflammation Ocular inflammation, including uveitis, retinitis and inflammation after surgery, is characterized by increased activity of PLA2. Therefore, UG can be administered topically, intraocularly or systemically to reduce ocular inflammation.

Arteriesclerosis Arteriosclerosis is a fibrotic thickening of the blood vessels throughout the body. It starts and / or is mediated by the deposit of Fn in the walls of the vasculature. Atherosclerosis is a form of arteriosclerosis that includes the deposit of cholesterol, in addition to the deposit of Fn. Therefore, UG can be administered to prevent or reduce arteriosclerosis.

Acute renal failure Acute renal failure (ARF) is commonly a consequence of remote organ inflammation, infection or direct trauma, which results in the release and activation of soluble PLA2 in the circulation. The damage to the kidneys during the IRA can be very severe, with acute tissue damage favored by the inflammation resolving into kidney fibrosis, giving rise to reduced kidney function in the long term. The anti-inflammatory and anti-fibrotic properties of UG are particularly important in the kidney as demonstrated by the KO UG mouse. The preferred route of administration is by injection or by systemic administration. In general, the following non-limiting list of conditions are those associated with the inhibition of PLA2 and / or the deposition of fibronectin, each of which are candidates for treatment or prevention by the method of the present invention: Articulation / bone: rheumatoid arthritis; Autoimmune: rheumatoid arthritis, multiple sclerosis, type I diabetes, uveitis, psoriasis, systemic lupus erythematosus (SLE), and Crohn's disease; Pancreas: pancreatitis; Peritoneum: peritonitis, appendicitis; Vascular / systemic: septic shock; vascular collagen disease, arteriosclerosis, atherosclerosis, anaphylactic shock, schistosomiasis and shock induced by trauma; Renal acute renal failure, bacterial infection of the kidneys, inflammation due to renal tumors, prevention of fibrosis resulting from chemotherapy or antibody therapy, prevention of diabetic nephropathy, prevention and / or treatment of idiopathic nephropathy; Liver: hepatitis, viral hepatitis and cirrhosis; Vejija: cystitis, inflammation of the urethra, inflammation of the ureter and inflammation of the bladder as interstitial cystitis; Reproductive / sex vaginitis, inflamed cervix, female: pelvisitis, ovarian inflammation (salpingitis), endometriosis, vaginal candidiasis and inflammation or fibrosis of the fallopian tubes; Reproductive / sex penis inflammation, male inflammation: the prostate, inflammation of the tubules and seminal vesicles, testicular inflammation and inflammation of the vas deferens, the epididy and the prostate gland; Ocular: uveitis, retinitis, trauma, damage from burns caused by chemicals or smoke, ocular inflammation due to CMV retinitis, conjunctivitis (bacterial infection), viral infection, ocular inflammation due to infectious agents, ocular inflammation after eye surgery, including cataract removal, laser surgery, corneal transplant, tumor removal, ocular inflammation due to retinoplastoma (tumor), ocular inflammation due to radiation exposure, inflammation due to allergic response; Heart: endocarditis Lungs: bronchial asthma, ARDS, pneumonia, idiopathic pulmonary fibrosis, pulmonary fibrosis resulting from chemotherapy (bleomycin, methotrexate), pulmonary fibrosis resulting from exposure to environmental chemicals (asbestos, cleaning fluids, contaminants, for example dioxin and PCB) in car exhaust gases), smoke inhalation, lung inflammation due to recovery from drowning, and neonatal SPR; Bowel: inflamed bowel disease, colitis, Crohn's disease, directiculitis, neonatal necrotizing enterocolitis, inflammation due to an infectious agent, rotavirus, poliovirus, HIV, gastric ulcers, gastroesophageal reflux disease and tonsillitis; Hemorrhoids; Transplants: administration after transplant surgery for any organ or tissue in order to control inflammation or fibrosis and rejections; Ears: otitis media, and Skin: psoriasis, urticaria, allergic and dermatitis, scleroderma, contact dermatitis, chemical dermatitis (due to poison ivy, poison oak and exposure to chemicals such as PCBs, chlorine, ammonia (cleaning agents / toxic agents) In addition, UG can be administered alone or in combination with other active agents or compositions commonly used in the treatment or prevention of conditions previously identified. These active agents or compositions include, but are not limited to, spheroids, non-spheroidal anti-inflammatories (NSAIDs), chemotherapeutic agents, analgesics, immunotherapeutics, antiviral agents, antifungal agents, vaccines, immunosuppressants, hemato-peptic growth factors, hormones, cytokines, antibodies, antithrombotic drugs, cardiovascular medications or fertility drugs. Also included are the oral tolerance drugs, vitamins and minerals. Moreover, UG can be administered as a carrier for the suppression of inflammation at the site of injection or administration of another therapeutic or prophylactic agent to suppress inflammation or an immune response caused by this agent. In this sense, the UG would allow the administration of an effective amount of the active agent for the objective indication. UG can also be administered as a cosmetic composition to limit fibrosis, the formation of scars or keloids resulting from wounds, or during a dermal inflammatory response. In addition, UG can be used as a blood supplement, that is, as an additive for donated human or synthetic blood. Finally, the UG can be used to improve the speed of artificial insemination by adding it to sperm in in vitro fertilized eggs and embryos before transferring them to the maternal womb in humans or other mammals. The present invention relates to the use of UG in the prevention or treatment of the conditions associated with PLA2 and fibronectin. With respect to the prevention of a disease state, "prevention" refers to preventing the development of the disease in a susceptible or potentially susceptible population, or limiting its severity or progress, while the term "treatment" refers to the relief of a disease or pathological condition. UG can be administered intravenously, or in the case of treatment of neonatal SPR / PPD and ARDS in adults, in the form of a liquid or semi-aerosol via the intratracheal tube. Other possible routes of administration include topical, ocular, dermal, transdermal, anal, systemic, intramuscular, slow release, oral, vaginal, intraduodenal, intraperitoneal and intracolonic. These compositions can be administered to an individual or patient in need of this administration in doses and by techniques well known to those skilled in the medical, nutritional or veterinary techniques taking into account factors such as age, sex, weight and condition of the patient. individual or specific patient, and the route of administration. The compositions of the present invention can also be administered in a controlled release formulation. The compositions can be co-administered or administered in sequence with other active agents, again taking into account factors such as the age, sex, weight and condition of the specific individual or patient, and the route of administration. Examples of compositions of the invention include edible compositions for oral administration as solid or liquid formulations, for example, capsules, tablets, pills and the like, liquid orifice preparations, eg, oral, nasal, anal, vaginal, etc., formulation as suspensions, syrups or elixirs; and, preparations for parenteral, subcutaneous, intradermal, intradermal, intramuscular or intravenous administration (e.g., injectable administration), such as sterile suspensions or emulsions. However, the active ingredient in the compositions can complex with the proteins so that, when administered to the bloodstream, coagulation can occur due to the precipitation of the blood proteins; and, the person skilled in the art should take this into consideration. In these compositions, the UG can be mixed with a suitable carrier, diluent or excipient such as sterile water, physiological saline, glucose, DMSO, ethanol or the like. The UG can be provided in a lyophilized for reconstitution, for example, in isotonic aqueous, saline, glucose or DMSO buffer. In certain saline solutions precipitation of rhUG has been observed; and this observation can be employed as a means to isolate compounds from the inventive, for example, by a "desalting" process. In addition, the invention also comprises a kit wherein the UG is provided. The kit may include a separate container containing a suitable carrier, diluent or excipient. The kit may include an additional agent that reduces or alleviates the effects of the conditions previously identified for co-administration or administration in sequence. The additional agent (s) may be provided in separate vessel (s) or in admixture with the UG. In addition, the kit may include instructions for mixing and combining the ingredients and / or administration. The invention also contemplates a method for the treatment or prevention of an inflammatory or fibrotic condition characterized by an insufficiency of functional endogenous UG, which consists in administering to a patient in need of this treatment a compensating amount of UG. The term "compensatory amount" means a quantity of UG necessary to bring the local or systemic pulmonary concentration of the total UG (functional UG, endogenous and exogenous UG) up to its normal range. More specifically, the normal range for the local lung concentration of the endogenous UG is approximately > 50 micrograms of UG / milligram of albumin or > 50 micrograms / liter. The normal range for serum UG concentration is > 15 micrograms / liter. The compositions of the invention contain native and / or recombinant hUG in an amount effective to achieve the intended purpose, namely, an increase in UG levels in plasma or tissue to produce the desired effect of selective inhibition of PLA2 and reduced inflammation. and / or the binding of fibronectin to mitigate its aggregation and / or deposition in fibrotic conditions. The compositions contain an effective amount of substantially pure native and / or recombinant human UG in association with a pharmaceutically acceptable carrier or diluent. In another aspect, the compositions of the invention contain an effective amount of native and / or recombinant hUG and lung surfactant, in association with a pharmaceutically acceptable carrier or diluent. Local intratracheal administration of UG to the lungs, sufficient for the inhibition of PLA2 activity and / or fibronectin deposition, requires practically pure amounts of UG in the range of 0.2 μg / kg to 500 mg / kg of protein in a single multiple dose or dose. UG is usually administered in a single-pole amount of 20 ng / kg up to 500 mg / kg, in a single dose or multiple doses, or as a continuous infusion of up to 10 grams. The native and / or recombinant hUG can also be administered together with artificial lung surfactant, such as Survanta, through the intra-tracheal route. The UG and Survanta (5 ml / kg) are co-administered and the UG does not bind to the surfactant, avoiding therapeutic functioning. The lung surfactant is generally present in the composition in an amount of about 10-90% by weight, more commonly about 20-80% by weight. The surfactant, by virtue of its very low surface tension, disperses on the inner surface of the lungs, carrying with it the UG. Systemic administration of UG via intravenous injection, sufficient for the inhibition of PLA2 activity and / or fibronectin deposition, requires quantities of substantially pure native and / or recombinant hUG in the range of 0.5 μg for a continuous infusion of several grams of protein for a prolonged period (days). Suitable formulations for injection and intratracheal delivery in semiaerosol include aqueous solutions of native and / or recombinant hUG optionally with viscosity modifiers and stabilizers. The term "PLA2 inhibitory effective amount" as is used herein, means the amount of UG that inhibits the activity of PLA2 and that reduces or alleviates inflammation in the patient's tissue or body- The term "effective amount that binds to fibronectin" means that amount of UG that it binds to fibronectin to reduce aggregation and / or deposition thereof, and prevents or reduces fibrillogenesis or fibrosis. The term "anti-inflammatory amount" as used herein means the amount that reduces or alleviates inflammation in the tissue or body. Usually, the amount of UG administered to adults for the treatment of inflammatory and fibrotic conditions will be individual boluses of 0.2 μg / kg up to 500 mg / kg or up to a few grams administered for a prolonged time. For neonates, in the treatment of neonatal MMR, the range will usually be 50 nanograms / kg up to 100 mg / kg in individual boluses or up to 10 grams administered continuously for a long time. The effective and safe rates of continuous infusion are between 50 ng / kg / hour up to 500 mg / kg / hour.

EXAMPLES The invention will now be described with reference to the following non-limiting examples. The parts and percentages are by weight unless stated otherwise.

Example 1: Live Experiments The recombinant human UG was obtained by the method of Mantile et al. (1993). One male and one female of the P. cyanocephalus species, weighing approximately 400 grams each, were removed by section C at 142 days of gestation. This is an established model of SPR (Coalson, J.J. et al., Baboon Model of BPD, II: Pathologic features, Exp. Mol. Pathol., 37: 355-350 (1982)). After delivery, the neonates were anesthetized with ketamine (10 mg / kg) and intubated with a 2.5 mm diameter endotracheal tube. Gases and blood pressure were monitored through an arterial line placed by percutaneous injection into the radial artery. A deep venous line was placed percutaneously into the saphenous vein through which fluids, antibiotics and medications were administered. The animals were kept in incubators heated with infrared, servo controlled and ventilated with a pressure controlled fan, with time cycle, standard, with humidifiers, at a temperature of 36-37 ° C. The initial parameters were Fi02 1.0, speed 40 / min, rate I / E 1: 1.5, positive end expiration pressure (PHEP) at 4 cm H20, maximum inspiratory pressure (PIP) as required to adapt the excursion of the chest. The Fi02 to 1.0 and the PIP was regulated to maintain the PaC02 at 40 ± 10 torr. Blood gases, hematocrit, electrolytes, prothrombin time, partial thromboplastin time and dextrostix [src] were monitored every hour. The blood extracted for the studies was replaced volumetrically with heparinized adult mandrel blood. The intravenous fluids were administered with electrolytes at 10 cc / kg / h and were increased as necessary when the heart rate exceeded 180 beats / minute. Sodium bicarbonate (2 meq / kg) was administered when the base deficit exceeded -10. Ampicillin (50 mg / kg / day in two divided doses) and Gentamicin (5 mg / kg / day in two divided doses) were administered continuously during the course of the experiment. One animal received surfactant plus PBS (treatment # 1), and the second animal (treatment # 2) received surfactant plus two doses of 1 mg / kg of rhUG. Both the surfactant and the rhUG were administered directly to the lungs through the endotracheal tube. The surfactant used was Survanta (Ross Labs), a surfactant preparation obtained from bovine lung tissue, with an apoprotein content of surfactant B and C in addition to phospholipids. The first dose of rhUG was administered with the surfactant and the second dose administered 4 hours after the first. The animals were analyzed for the gases blood arteries, electrolytes and ECG. These were sacrificed 50 hours after the start of the surfactant therapy. The lungs were washed at 24 and 48 hours with PBS containing protease inhibitors (PMSF, 10 μg / ml leupeptin, 10 μg / ml pepstatin and bacitracin). These were frozen at -80 ° C until assayed for PLAz activity. The total proteins were determined by the Bradford method (BioRad). The activity of PLA2 in lung washes was measured according to Levin et al. (1986, supra) and are presented in the following Table.

Table 3. Results of the In Vivo test of the UG The data provided before is the average of two determinations. The results show that endotracheal administration of rhUG inhibits PLA2 in vivo. The animals that received surfactant and rhUG had an appreciably lower activity of PLA2 in their Lung wash fluid compared to animals that received surfactant without rhUG. The data confirm that the administration of rhUG together with the surfactant is beneficial in the protection of the phospholipids of the surfactant.

Example 2: Inhibition of hydrolysis of artificial surfactant by soluble PLA2 in vitro. RhUG inhibits the hydrolysis of artificial surfactant by soluble PLA2 in vi tro. Survanta is an artificial surfactant from the bovine lung and is used to treat preterm infants with SPR and adults with SPR (SPRA). Hydrolysis of Survanta by soluble PLA2, Group 1, ie porcine pancreatic PLA2 (Boehringer Mannheim) is characterized by its ability to compete as a substrate with a fluorescent phosphatidyl choline substrate (Cayman Chemicals), generating arachidonic acid as a product. Survanta is a substrate for in vitro degradation by soluble PLA2 from Group I. Survanta is rapidly degraded in vi tro by the PLA2 found in the extracellular fluids of a human lung. RhUG inhibits the degradation of Survanta in vi tro.

Example 3: Construction of a mouse knocked out with UG A transgenic KO UG mouse was created for the purpose of determining the function of uteroglobin in the physiology of mammals, as well as to treat a model for UG as a therapeutic in various inflammatory clinical conditions. The first step was to construct a suitable DNA vector with which to direct and interrupt the murine endogenous uteroglobin gene. The 3.2 kb BamHI-EcoRI DNA fragment containing exon 3 and the flanking sequences of the uteroglobin gene from mouse strain 129 / SVJ (Ray, 1993) were subcloned into the corresponding sites of the pPNW vector as described in FIG. Lei et al (1996). A 0.9 kb fragment containing part of exon 2 and its upstream sequence was amplified by PCR with Primer-L primers (from Intron 1): 5 '-TTC CA GGC AGA ACÁ TTT GAG AC-3'; Primer-R (from Exon 2): 5 '-TCT GAG CCA GGG TTG AAA GG C-3' with the Notl and Xhol restriction sites designed in terms for directional subcloning in the vector targeting the gene. In this construction, 79 bp of Exon 2 was deleted, which codes for 27 amino acids. The PCR fragment was placed upstream of the gene coding for neomycin resistance in pPNW, generating the vector that targets the gene, pPNWUG. The vector is shown in Figure 2A, in which the PGK-neo cassette interrupts the uteroglobin, breaking the coding sequence of the protein. The vector that targets the pPNWUG gene was linearized with Notl and electroporated into ES cells Ri according to Nagy, A., al. PNAS 90: 8424 (1993). Giantclovir and the G-418 selection of electroporated cells produced 156 clones. Southern blot analysis (DNA) identified a 5.1 kb HindIII fragment of wild type uteroglobin allele and an additional 8.2 kb HindIII fragment resulting from homologous recombination in 3 of the 156 clones, shown in Figure 2B. These ES Rl clones were injected into the C57BL / 6 blastocysts according to Capecchi, Science 244: 1288 (1989). Two different lines of mice were produced that descended from different chimeric founders. The heterozygous progeny (UG + ~) carrying the locus of the directed uteroglobin gene were paired and the progeny genotypes were analyzed by PCR as shown in Figure 2C, as well as by Southern blot, as shown in Figure 2D.

Example 4: Verification of the gene knockout for UG and the absence of the murine UG protein (mUG) To verify that the homozygous knockout mouse (UG - / - \ does not possess any detectable mUG, mice targeted with the gene for uteroglobin they were tested for the expression of UG-mRNA and the mUG protein in various organs that includes the lungs. An experimental protocol was approved by the Institutional Animal Care and the use committee. Total RNAs were isolated from different organs of UG + +, UG + ~ and UG ~ _ mice. The reverse transcribed polymerase chain reaction (RT-PCR) was used to detect mUG-mRNA. The target molecules were reverse transcribed using the primer specific for mUG, mPr (5 '-ATC TTG CTT ACA CAG AGG ACT TG-3'), and the generated cDNA was amplified using the mPr and mPl PCR primers (5 '- ATC GCC ATC ACÁ ATC ACT GT-3 '). The PCR product was hybridized with an oligonucleotide probe, mPp (5 '-ATC AGA GTC TGG TTA TFT GGC ATC C-3') from exon-2 of the gene sequence for UG. The primers and probe used in the RT-PCR, mouse GAPDH are as follows; mGAPDH-r (5 '-GGC ATC GAA GGT GGA AGA GT-3'); mGAPDH-1 (5 '-ATG GCC TTC CGT GTT CCT AC-3'); mGAPDH-p (5 '-GAA GGT GGT GAA GCA GGC ATC TGA GG-3") Figure 2E shows that mUG-mRNA was detected in the lungs of UG + + and UG + /", but not in mice UG_ / " Similar data (not shown) demonstrate that mUG-mRNA is not present in the prostate or uterus of UG_ / ~ mice, but is present in mice with an intact uteroglobin gene, Lp, immunoprecipitation and Western blot analysis of the mUG protein in the lungs produced results corroborated similar, as shown in Figure 2F. Lysates of kidneys, liver and lung tissue of mice UG + + and UG_ ~ were prepared by homogenizing in a buffer (10 mM Tris-HCl, pH 7.5, 1% Triton X-100, 0.2% deoxycholate, 150 mM NaCl, 5 mM EDTA) with a 2 mM content of phenylmethylsulfonyl fluoride and 20 μg / ml of aprotinin, leupeptin and pepstatin A each. The homogenates were centrifuged at 17,500 xg for 30 minutes at 4 ° C and immunoprecipitates as described (E. Harlow and D. Lane, Antibodies, a laboratory manual, the Cold Spring Harbor Labor Press, Cold Spring Harbor, NY Ed. , 1988) by incubation of tissue lysates or plasma proteins (1 mg / mL) with rabbit antibody against murine Fn (1: 100 dilution). Co-immunoprecipitation of purified murine Fn (mFn) and rhUG (Mantile, G, et al., J. Biol. Chem. 267: 20343 (1993)) was performed by incubating equimolar concentrations of mFn with rhUG in the presence of 10% glycerol, Tris-HCl, 50mM, pH 7.5, 250 M NaCl, 4.3 mM sodium phosphate at 4 ° C for 1 hour. Followed by the addition of anti-mFn antibody (1: 100 dilution). Equal amounts of tissue proteins extracted (30 μg) or immunoprecipitated were resolved on SDS-polyacrylamide gels 4-20% or 6% under reducing conditions, followed by Western blot analysis with rabbit antibodies against murine Fn (dilution 1 : 2000) or UG (dilution 1: 2000). No mUG was detected in tissues or fluids from UG _ / _ mice, whereas tissues from UG + / + and UG + ~ mice contained the mUG protein. Finally, the histopathological analysis of the lungs of UG_ "mice only / lacked specific immunotinization of mUG in bronchiolar epithelial cells.The lung tissues of UG_ / ~, UG +" and UG + + mice were fixed in Bouin's fluid or in 10% formaldehyde fixer in neutral buffer, embedded in paraffin and sectioned in 4-6 microns. These were stained with hemotoxilin [sic] and eosin (H &E). The selected tissues were stained by the Masson's trichrome method for the detection of collagen, PTAH for fibrin or Congo red for the amyloid protein. For the immunohistochemical detection of mUG and mFn, the Vectastain rabbit Elite ABC kit (Vector Laboratories) was used. The rabbit antibody (Cytlmmune) for mUG was increased using a synthetic peptide (Peptide Technologies, Inc.) corresponding to the amino acid sequence for mUG (Lys28 to Thr49, specifically KPFNPGSDLQNAGTQLKRLVDT). The rabbit antibody for mFn (GIBCO BRL) was used at a dilution of 1: 1000, and the antibody for mUG was used at 1: 500. These three series of results confirm that the homozygous mouse knocked out with uteroglobin, UG- / ~, lacks the mUG protein or any detectable piece of protein.

Example 5: Mouse phenotype knocked out with uteroglobin Of the 179 mice born for crosses of UG + ~ mice, 46 (26%) were + / +, 90 (50%) of the +/- and 43 (24%) of the genotype UG "7", indicating that the interrupted mUG locus is inherited in a Mendelian form and that the UG + / +, UG + / ~ and UG - / ~ mice were equally viable at birth. However, the UG _ ~ mice presented a novel phenotype in which they developed a progressive disease characterized by cachexia, strong proteinuria and hypocalcemia associated with deep weight loss. Proteinuria is a state in which abnormally high levels of albumin and other whey proteins are excreted in the urine. This is indicative of glomerular dysfunction and renal failure. The histopathological examination of the kidneys of affected animals (as described above for the lungs) showed the fulminating renal glomerular disease shown in Figure 3. Compared with the glomeruli of the UG + / + mice, those of the UG mice " were cellular type and had massive eosinophilic proteinaceous deposits The time of fatal renal disease in UG - / - mice was initiated at the beginning (period of 4-5 weeks) started at the end (period of 10 months) Those mice UG - / - that Initially they seemed healthy at 4 weeks of age they had focal glomerular deposits at two months of age. At about 10 months these mice had extreme cachexia similar to that of the dying mice of the early onset of the disease. The heterozygotes had a more moderate form of kidney disease observed in the UG_ / ~ mice. The histopathology of the kidneys of mice with late-onset disease showed not only severe glomerulopathy as in the disease initiated at the beginning, but also had marked fibrosis of the renal parenchyma and tubular hyperplasia (see Figure 3). Although the predominant pathology in UG_ ~ mice was found in the kidneys, histopathological studies also discovered occasional focal areas of necrosis in the pancreas that appeared to have vascular orientation. In addition, focal areas were also found in the thymus and in the splenic structures that suggest apoptotic bodies. It is interesting to note that the pancreas expresses the gene for mUG and this organ is also a rich source of extracellular PLA2 from group I; Since this is mainly a digestive enzyme, its action can cause tissue injury. Because uteroglobin proteins have been reported with immunomodulatory and anti-inflammatory properties and because it is known that amyloidosis Reactive reaction occurs in response to inflammation, it is very likely that glomerular deposits in null mice in mUG were amyloid proteins. Reactive amyloidosis is characterized by the deposition of amyloid protein and immune complexes. The identity of renal deposits in UG ~ / _ mice was established by immunohistochemistry of kidney sections. Kidney sections from UG_ ~ and UG + + mice were stained with Congo red and examined under polarized light. Amyloid proteins produce a positive double refraction in this test. However, the glomeruli of the UG "_ mice were clearly negative, and immunofluorescence studies for the presence of IgA, IgG or IgM immunocomplexes in the glomeruli of UG_ /" mice and immunohistochemical analyzes for the presence of major amyloid proteins were also negative. Thus, the glomerular deposits of the UG ~ _ mice did not contain amyloid proteins or immunocomplexes and, therefore, do not appear to be the result of an inflammatory response.

Example 6: Detection of Fn and collagen in UG kidneys ~ / - Kidney deposits of UG_ ~ mice were examined by transmission electron microscopy to elucidate their structure and morphology. The UG- / ~ mouse kidney with glomerular lesion was fixed in formalin and embedded in resin Epoxy The thin sections were stained with uranyl acetate and lead citrate for examination under the electron microscope. The photomicrographs were taken at 6000X or at 60,000 X. The deposits mainly contained two types of fibrillar structures: one type of long and striated fibrils that are relatively infrequent, the other short and diffuse fibrils that are more abundant (Figures 3E and 3F). Because ECM proteins, such as collagen and fibronectin, produce similar fibrillar structures, glomerular deposits in UG_ _ mice can contain these proteins. The glomerular deposits were then analyzed by immunofluorescence using anti-mFn antibodies. The sections of formalin-fixed tissues were used for immunofluorescence using goat anti-rabbit IgG, conjugated with FITC and rabbit anti-mFn. In the same way, immunofluorescence studies were also carried out using antibodies specific for mFn, collagen I and 111 / vitronectin, laminin and osteopontin. The epifluorescence was photographed using a Zeiss Axiophot microscope. Fn-specific immunofluorescence in the renal glomeruli of wild-type mice was virtually undetectable (Figure 3G), compared to the glomeruli of the littermates UG ~ / _ which was more intense (Figure 3H). When the Masson's trichrome stain was used, the glomeruli of the UG + / + mice were negative (Figure 31) and those of UG ~ / _ (Figure 3J) were positive, suggesting the presence of collagen in the glomerular deposits. Immunofluorescence, using antibodies specific for collagen I and collagen III, confirmed these results. Because it is known that Fn interacts with other extracellular matrix (ECM) proteins, we also tested the presence of laminin, vitronectin and osteopontin in the glomeruli of UH + + and UG- / ~ mice by immunohistochemistry, the results of which were negative Example 7: Kidneys from UG- ~ mice do not produce excess Fn. To determine if the excess production of Fn can represent its deposit in the renal glomeruli, we tested the relative amount of Fn-mRNA in the kidneys, lungs and liver of UG mice ~ _ and UG + + by RT-PCR and densitometry. The results indicate that, relative amounts of Fn-mRNA were practically identical in the animals UG + + and UG_ / ". Thus, the overproduction of Fn-mRNA was not a probable cause of Fn deposition in the glomeruli of UG mice" We then compared the Fn protein in the plasma, kidneys and liver of UG-UG + / + mice by SDS-PAGE under reducing conditions and Western blot. In the plasma, the kidneys and the liver of wild type mice only species of Fn of 220 kD could be detected; however, although the plasma and the liver lysate of UG_ ~ mice had the Fn band of 220 kD, the kidney lysates contained another band of multimeric Fn, with a different covalent bond (Figure 4A).

Example 8: Elevated serum PLA2 activity in UG- ~ mice Based on current concepts, critical initial steps in the assembly to the Fn matrix and fibrinogenesis, at least on the cell surface is considered to include integrin activation and Fn autoaggregation. Because UG is a potent inhibitor of soluble phospholipase A2 (sPLA2) / a key enzyme in the inflammatory pathway, the lack of mUG in UG_ ~ mice may contribute to the development of glomerulonephritis and inflammatory kidney disease. Thus, we measured the activity of PLA2 in the serum of UG + / + mice (n = 3 = and UG "/ _ (n = 3) of the same age, sex and weight, the animals were sacrificed and PLA2 activities in serum of each sample were measured in triplicate using the PLA2 assay kit (Caymen Chemical) according to the manufacturer's instructions.The concentrations of the protein in the sera were determined by the Bradford assay (Bio Rad) and were calculated the specific activities of PLA2.The specific activities (μmol / min / mg protein) of the PLA2 in serum of the UG- / "[36 + 3.3 (SEM)] mice were significantly higher (p <0.05) than those of the UG + / + mice [18 + 2.5 (SEM)]. These results increased the possibility that the activity of increased PLA2 may give rise to increased production of lysophosphatidic acid (LPA) and consequently favor integrin activation and autoaggregation of Fn in UG_ ~ mice.

Example 9: Interaction of uteroglobin and fibronectin in vitro To understand more how uteroglobin can prevent self-assembly of Fn, the ability of rhUG to interrupt the mFn-Fn interaction in vitro was determined. Equimolar concentrations of rhUG and mFn were incubated to allow any binding to protein or other interactions, then immunoprecipitated with anit-Fn antibody, and the immunoprecipitates were resolved by SDS-PAGE under reducing conditions. Western blot analysis, as already described, with mFn or mUG antibody detected each protein, respectively. The results show this co-immunoprecipitated fibronectin with rhUG (Figure 4B). To confirm these results, the 125I-rhUG was incubated with mFn and the complexes were resolved by electrophoresis using a 6% polyacrylamide gel under non-denaturing and nonreducing conditions (Figure 4C). The Detection of an Fn-UG heteromer in an autoradiogram (lane 2) showed these interactions of soluble Fn with UG in vitro. To assess whether the heteromerization of Fn-UG is carried out in vivo, the plasma of UG + / + and UG ~ / _ mice was immunoprecipitated with an anti-mFn antibody that does not cross-react with rhUG (Figure 4D). The anti-mFn antibody co-precipitated the mFn and rhUG from the plasma of the mice UG + / + but not from UG mice, suggesting that Fn-UG heteromers are present in the plasma of UG + + mice. Therefore, the Fn-UG complex is not simply an artifact formed in vitro but occurs naturally in serum. To determine the specificity and affinity of the binding of UG to the Fn, we incubated 125I-Fn with the unlabeled Fn in the presence and absence of UG. Any of the complexes were crosslinked by affinity with disuccinimidyl suberate (DSS). Using 24-well plates coated with human Fn (hFn) (Collaborative Biomedical Products), 3 μl of 125 I-Fn (Sp. Act. 6 mCi / mg:? CN Biomedicals) were incubated in the absence and presence of UG or Fn (10 ~ 12-10 ~ 6 M) in 500 μl of HBSS at room temperature for 2 hours. SDS-PAGE and western blot analysis of all Fn antibodies with UG failed to detect any contamination with UG. The radiolabeled complex was washed twice with PBS, solubilized in IN NaOH, neutralized with IN HCl, and the radioactivity was measured by a gamma counter. In a separate experiment, 125 I-hFn (3 μl) was incubated with 20 μl (1 mg / ml) of mouse Fn in 40 μl of HBSS, pH 7.6, in the absence or presence of increasing concentrations of reduced rhUG (5-500). μg) at room temperature for two hours. The samples were cross-linked with 0.20 M DSS at room temperature for 20 minutes, boiled in SDS-sample buffer for 5 minutes, subjected to electrophoresis on 4-20% SDS-polyacrylamide gel and subjected to autoradiography. In the absence of UG, 125I-Fn formed a high molecular weight radioactive complex with unlabeled Fn, but in the presence of UG the formation of Fn-Fn aggregates was inhibited in a UG-dependent manner (Figure 4E). To determine if there is any difference between the binding affinities of Fn by UG and of this Fn by itself, binding experiments were performed in which 1251-Fn was incubated with unlabeled Fn and immobilized on multiple well plates along with variable concentrations of UG. In separate experiments, binding studies of 125I-Fn with unlabeled Fn, immobilized using various concentrations of soluble, unlabeled Fn, were also performed. Scatchard analyzes of the data from both types of binding experiments yielded straight lines with dissociation constants (kds) of 13 nM for binding from UG to Fn and 176 nM for the union of Fn itself. These results suggest that, due to a relatively high binding affinity of the UG for Fn, the UG effectively counteracts the autoaggregation of the Fn. The affinity-crosslinking experiments in which the radioiodinated collagen I- (1 5I) was incubated with unlabeled Fn in the absence or presence of UG, were also performed as already described for Fn. 15 μl of denatured or undenatured 125 I-collagen I (Sp. Act. 65.4 mCi / mg) were incubated with Fn in the presence or absence of reduced (250 μg) UG, cross-linked by affinity, subjected to electrophoresis and autoradiography. The results indicate that the UG counteracts the formation of 125 I-collagen-Fn aggregates of high molecular weight (Figure 4F).

Example 10: In vivo inhibition of glomerular hFn deposition by rhUG To test whether rhUG protects the renal glomeruli from accumulation of Fn, soluble human Fn (hFn), alone or hFn mixed with equimolar concentrations of rhUG was administered intravenously to litters of UG + / + mice and UG_ / ~ apparently healthy. Human Fn (500 μg / 150 μl PBS) was administered in the tail vein of UG + / + and UG mice apparently healthy at 2 months of age, approximately 22 g.

In the same manner, the control mice were injected with a mixture of 500 μg of hFn with equimolar concentrations of rhUG or albumin in 150 μl of PBS. Twenty-four hours after the last injection, the mice were sacrificed and different organs were fixed in formalin with buffer. The histological sections of the kidneys and other organs were examined by immunofluorescence with a monospecific anti-hFn antibody (GIBCO BRL, clone 19 and FITC-conjugated rabbit anti-mouse IgG (Cappel).) In a separate experiment, the UG + + mice were injected with 1 mg of hFn alone in 150 μl of PBS per day for 3 consecutive days The reason for injecting human Fn was to be able to discriminate between the murine, endogenous Fn and the administered hFn.The method of intravenous administration and detection by immunohistochemistry The immunofluorescence of human Fn in the glomeruli of UG + / + wild-type mice injected with a mixture of hFn and rhUG (molar ratio 1: 1) or with hFn alone was similar (Figures 5A and 5B). However, UG_ / "mice injected with a mixture of hFn and UG showed little specific immunofluorescence of hFn in the glomeruli (Figure 5C), while those who received Fn alone presented immunofluorescence with greater intensity (Figure D) . The administration of a mixture of hFn and BSA, as a control, did not produce a protective effect. To determine if this UG protective effector could be overcome by injecting greater amounts of Fn in UG + / + mice, we injected 1 mg of hFn per animal daily for 3 consecutive days. Although intravenous administration of hFn to UG + + mice at lower doses (500 μg / animal) was not effective in causing any appreciable glomerular deposition (Figure 5A), administration of higher doses (3 mg / animal) caused a significant accumulation. In this way, UG prevents the deposition of glomerular Fn and UG + / + mice, contrary to UG ~ / _ mice, have a higher threshold for the accumulation of soluble Fn, due to the presence of endogenous UG.

Example 11: Inhibition of fibrinogenesis and assembly to the Fn matrix by rhUG in tissue culture cells To determine whether UG prevents fibrinogenesis by Fn and assembly to the matrix in a common in vitro tissue culture assay Mouse embryonic fibroblasts were cultured in medium containing soluble hFn alone or a mixture of equimolar concentrations of hFn and rhUG. The assembly to the Fn matrix and fibrillogenesis in cultured cells (CRL6336, ATCC) were determined as described. The level of fibrillogenesis observed in the Cells from cultures treated with hFn alone were much larger (Figure 5E) compared to those that received a mixture of hFn and rhUG (Figure 5F).

Example 12: Detection of UG-Fn complexes in clinical samples The detection of UG-Fn complexes in clinical samples of bodily fluids such as serum, BAL fluids and sputum is important in determining the function of this complex in human diseases. A solution phase diagnostic assay was developed for the detection of the UG-Fn complexes and the assay format is shown in Figure 6. The capture antibody, covalently linked to a solid support, is a polyclonal rabbit , monospecific, grown against human protein. The solid support can be a bead, such as a magnetic bead, a tube or an ELISA plate. The solid support produces the flexibility to perform wash steps after each binding reaction in order to obtain more consistent results with a variety of sample types. The detection antibody is specific for Fn and is available to some commercial sources. An anti-IgG antibody, conjugated to an enzyme such as horseradish peroxidase (HRP), is then used to detect anti-Fn IgG at the end of the molecular chain in a normal enzymatic reaction in which the substrate of the enzyme is converted into a chromogenic or fluorogenic compound that is quantified with a spectrophotometer or fluoride (Amersham). The limit of detection for this assay is 500 μg of the UG-Fn complex per ml of the sample fluid.

Example 13: Uteroglobin insufficiencies A transient but acute insufficiency of hUG is created by the blood cleansing technique known as clinical dialysis, including hemodialysis, peritoneal dialysis and continuous dialysis (CRRT). All forms of clinical dialysis include the use of a semipermeable membrane to filter out toxic, bodily waste products that include chemical metabolites such as urea and small proteins such as beta2-microglobulin, out of the blood. UG is an extremely compact protein, known for its anomalous migration in SDS-PAGE, corresponding to a molecular weight of approximately 10-13 kDa, despite its actual molecular weight of 15-7 kDa. Therefore, it was expected that the UG dimer would behave as a 10-13 kDa protein in dialysis experiments. Surprisingly, it was found that the dimer is so compact that it passes through a MWCO dialysis membrane of 8.0 kDa. The UG also passes through a MWCO dialysis membrane of 14.0 kDa. The composition of the dialysis membranes used in these examples are similar, if not identical, to the composition of most of the membranes manufactured and used for clinical dialysis. These consist of regenerated cellulose or cellulose acetate. For this experiment, 1.0 ml aliquots of two partially purified rhUG cell lysates (one> 90% pure and one approximately 70% pure), without buffer additives, were dialyzed against 1000 ml of 50 mM non-buffered ammonium acetate, using three sizes of dialysis tubing: 3.5 kDa, 8.0 kDa and 14.0 kDa (Spectra / Por, Thomas Scientific). There were four changes of buffer for each sample during a time of 48 hours, everything was done at room temperature (approximately 25-27 ° C). The appearance of each dialysis sample changed from a light yellow liquid to a clear, colorless liquid. The dialysis tube was checked for leaks at the beginning and end of the process by brief application of pressure directly to the pipeline (squeezing) and the observation of any leakage, of which there was none. The pipe was fastened double at each end to ensure there were no leaks. Figure 7 shows the SDS-PAGE analysis of these results. The sample before dialysis, 90% pure is shown in the 7th and 8th row followed by the three samples after dialysis in strips 1, 2 and 3. The UG dimer was no longer present in the bands representing the samples dialysed with 8.0 kDa MWCO membranes. These results were then confirmed with different batches of partially purified UG preparations. Although the invention has been described together with what is currently considered the most practical and preferred modalities, it will be understood that the invention is not limited to the described modalities but, on the contrary, it is proposed to cover the different modifications and equivalent arrangements included within of the spirit and scope of the appended claims.

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Claims (109)

1. A pharmaceutical composition containing an amount effective to inhibit PLA2 from recombinant human uteroglobin or a fragment or derivative thereof and a pharmaceutically acceptable carrier or diluent.

2. The pharmaceutical composition of claim 1 further contains a lung surfactant.

3. The pharmaceutical composition of claim 1, wherein the rhUG has a purity of about 75% to about 100%.

4. The pharmaceutical composition of claim 1, wherein the rhUG has a purity of about 90% to about 100%.

5. The pharmaceutical composition of claim 1, wherein the rhUG has a purity of at least 95%.

6. A pharmaceutical composition containing an effective amount that binds to fibronectin of a recombinant human uteroglobin (rhUG) or a fragment or a derivative thereof and a pharmaceutically acceptable carrier or diluent.

7. The pharmaceutical composition of claim 6 further contains a lung surfactant.

8. The pharmaceutical composition of claim 6, wherein the rhUG has a purity of about 75% at approximately 100.

9. The pharmaceutical composition of claim 6, wherein the rhUG has a purity of about 90% to about 100%.

10. The pharmaceutical composition of claim 6, wherein the rhUG has a purity of at least 95%.

11. A pharmaceutical composition containing an effective amount that inhibits PLAE O binds to UG fibronectin in admixture with an active agent for the treatment of an objective indication.

12. The pharmaceutical composition of claim 11, wherein the effective amount that inhibits PLA? or that binds to rhUG fibronectin reduces inflammation and thus ensures that an effective amount of the active agent reaches the treatment site.

13. A method for inhibiting PLA2 enzymes in vivo in a mammal in need of such treatment, the method consists of administering to the mammal an effective inhibitory amount of PLA? of recombinant native or human uteroglobin (UG) or a fragment or derivative thereof.

The method of claim 13, wherein the UG has a purity of between about 75% to about 100%.

15. The method of claim 13, wherein the UG has a purity of between about 90% a approximately 100%.

16. The method of claim 13, wherein the UG has a purity of at least 95%.

17. The method of claim 13, wherein the UG is recombinant (rhUG), having virtually the same sequence as the native human uteroglobin.

18. The method of claim 13, wherein the PLA2 inhibitory effective amount of UG is 20 mg / kg to 500 mg / kg.

The method of claim 13, wherein the UG is administered in association with a lung surfactant.

The method of claim 19, wherein the lung surfactant is present in an amount of about 20% to about 80% by weight.

The method of claim 13, wherein the UG is administered by the endotracheal, ophthalmic, intravenous, systemic, intraperitoneal, intramuscular or oral routes.

22. The method of claim 13, wherein the UG is administered as a liquid or semi-aerosol by an intratracheal tube.

The method of claim 13, wherein the method is for the treatment of a condition selected from the group consisting of systemic inflammation, asthma, cystic fibrosis, ocular inflammation, delivery prematurity, infertility, rheumatoid arthritis, type I diabetes, nephropathy, inflammatory bowel disease, Crohn's disease, ulcerative colitis, pancreatitis, peritonitis, allergy, multiple organ failure, ARDS, acute renal failure, inflammation secondary to infection or surgery and rejection of transplanted organ.

24- The method of claim 23, wherein the ocular inflammation results from autoimmune uveitis and corneal transplant surgery.

25. The method of claim 13, wherein the method is for the treatment of inflammation and immune modulation.

26. A method of treating or preventing an inflammatory condition in a patient in need of such treatment, the method comprising administering to the patient an anti-inflammatory effective amount of native or recombinant UG or a fragment or derivative thereof.

27. The method of claim 26, wherein the UG has a purity of between about 75% to about 100%.

28- The method of claim 26, wherein the UG has a purity of between about 90% to about 100%.

29. The method of claim 26, wherein the UG has a purity of at least 95%.

30. The method of claim 26, wherein the UG is recombinant (rhUG) and has virtually the same sequence as the native human uteroglobin.

31. The method of claim 26, wherein the amount effective to inhibit PLA2 from the UG is 20 mg / kg a 500 mg / kg.

32. The method of claim 26, wherein the UG is administered in association with a lung surfactant.

33. The method of claim 32, wherein the lung surfactant is present in an amount of about 20% to about 80% by weight.

34. The method of claim 26, wherein the UG is administered by the endotracheal, ophthalmic, intravenous, systemic, intraperitoneal, intramuscular or oral routes.

35. The method of claim 26, wherein the UG is administered as a liquid or semi-aerosol through an intratracheal tube.

36. The method of claim 26, wherein the inflammatory condition is neonatal SPR.

37. The method of claim 26, wherein the inflammatory condition is adult SPR.

38. The method of claim 26, wherein the inflammatory condition is caused by a viral infection, bacterial infection, parasitic infection or infection. fungal

39. The method of claim 26, wherein the method is for the treatment of an inflammatory condition selected from the group consisting of systemic inflammation, asthma, cystic fibrosis, ocular inflammation, premature delivery, multiple organ failure, rheumatoid arthritis, pancreatitis. , septic shock, collagen vascular disease, anaphylactic shock, shock induced by trauma, acute renal failure, cystitis, inflammation of the urethra, inflammation of the ureter, inflammation of the bladder, interstitial cystitis, vaginitis, inflamed cervix, pelvic inflammatory disease, inflammation of the ovariosalpingitis [sic], inflammation of the fallopian tubes, inflammation of the penis, inflammation of the prostate, inflammation of the seminal tubules and seminal vesicles, testicular inflammation, inflammation of the vas deferens, the epididymis and the prostate gland, retinitis, damage from burns due to fire or chemical burns, asthma bronchial, SPRA, neonatal SPR, direticulitis, neonatal necrotizing enterocolitis, gastric ulcers, gastrointestinal reflux disease, tonsillitis, hemorrhoids, otitis media, psoriasis, urticaria, allergic dermatitis, contact dermatitis, chemical dermatitis, endocarditis and infertility.

40. The method of claim 39, wherein the inflammation is ocular inflammation resulting from autoimmune uveitis, CMV retinitis, bacterial infection, viral infection, parasitic infection, the presence of an infectious agent, retinoblastoma, radiation exposure, allergic response and corneal transplant surgery.

41. A method for the treatment or prevention of a fibrotic condition in a patient in need of such treatment, the method comprising the step of administering to a patient an effective amount that binds to native or recombinant UG fibronectin or a fragment or derivative Of the same.

42. The method of claim 41, wherein the UG has a purity of between about 75% to about 100%.

43. The method of claim 41, wherein the UG has a purity of between about 90% to about 100%.

44. The method of claim 41, wherein the UG has a purity of at least 95%.

45. The method of claim 41, wherein the UG is recombinant (rhUG) and has virtually the same sequence as the native human uteroglobin.

46. The method of claim 41, wherein the effective amount of UG that binds fibronectin is 20 mg / kg to 500 mg / kg.

47. The method of claim 41, wherein the UGit is administered in association with a lung surfactant.

48. The method of claim 47, wherein the lung surfactant is present in an amount of about 20% to about 80% by weight.

49. The method of claim 41, wherein the UG is administered by the endotracheal, ophthalmic, intravenous, intraperitoneal, intramuscular, liquid or oral routes.

50. The method of claim 41, wherein the UG is administered as a liquid or semi-aerosol by an intratracheal tube.

51. The method of claim 41, wherein the fibrotic patient is pulmonary fibrosis.

52. The method of claim 41, wherein the fibrotic condition is renal fibrosis.

53. The method of claim 41, wherein the fibrotic condition is vascular fibrosis.

54. A method for the treatment or prevention of an inflammatory or fibrotic condition characterized by endogenous UG insufficiency, the method comprising administering to a patient in need of such treatment, a compensatory amount of native or recombinant UG or a fragment or derivative Of the same.

55. The method of claim 54, wherein the UG has a purity of between about 75% to about 100%.

56. The method of claim 54, wherein the UG has a purity of between about 90% to 100%.

57. The method of claim 54, wherein the UG has a purity of at least 95%.

58. The method of claim 54, wherein the UG is recombinant (rhUG) and has virtually the same sequence as the native human uteroglobin.

59. The method of claim 54, wherein the amount effective to inhibit PLA2 from the UG is 20 mg / kg a 500 mg / kg.

60. The method of claim 54, wherein the UG is administered in association with a lung surfactant.

61. The method of claim 60, wherein the lung surfactant is present in an amount of about 20% to about 80% by weight.

62. The method of claim 54, wherein the UG is administered by the endotracheal, ophthalmic, intravenous, systemic, intraperitoneal, intramuscular or oral routes.

63. The method of claim 54, wherein the UG is administered as a liquid or semi-aerosol through an intratracheal tube.

64. The method of claim 54, wherein the method is for the treatment of a condition inflammatory or fibrotic characterized by an insufficiency of endogenous UG, where the condition is selected from the group consisting of bronchopulmonary dysplasia, complications of hemodialysis, chronic obstructive pulmonary disease due to bleomycin and inherited glomerulopathy.

65. The method of claim 26, wherein the UG is administered in combination with an active agent selected from the group consisting of spheroids, non-spheroidal anti-inflammatory agents, chemotherapeutics, analgesics, antibodies, antiparasites, antivirals, antibiotics, antifungals, vaccines, vaccines against tumor, immunosuppressants, hematopoietic growth factors, antithrombotic, cardiovascular drugs, one or more vitamin and mineral supplements and fertility drugs.

66. The method of claim 44, wherein the UG is administered in combination with an active agent selected from the group consisting of spheroids, non-spheroidal anti-inflammatory agents, chemotherapeutics, analgesics, antibodies, antiparasites, antivirals, antibiotics, antifungals, vaccines, vaccines against tumor, immunosuppressants, growth factors, antithrombotics, cardiovascular drugs, one or more vitamin and mineral supplements and fertility drugs.

67. The method of claim 54, wherein the UG is administered in combination with an active agent selected from the group consisting of spheroids, non-spheroid anti-inflammatory agents, chemotherapeutics, analgesics, antibodies, antiparasitic, antivirals, antibiotics, antifungals, vaccines, vaccines against tumor, immunosuppressants, growth factors, antithrombotics, cardiovascular drugs, one or more vitamin and mineral supplements and fertility drugs.

68. A cosmetic composition containing an amount effective to inhibit PLA2 from native or recombinant human uteroglobin (UG) or a fragment or derivative thereof and a pharmaceutically acceptable diluent carrier.

69. The cosmetic composition of claim 68 further contains a lung surfactant.

70. The cosmetic composition of claim 68, wherein the UG has a purity of about 75% to about 100%.

71. The cosmetic composition of claim 68, wherein the UG has a purity of about 90% to about 100%.

72. The cosmetic composition of claim 68, wherein the UG has a purity of at least 95%.

73. A cosmetic composition containing an effective amount that binds human uteroglobin fibronectin native or recombinant (UG) a fragment or derivative thereof, and an acceptable carrier or diluent.

74. The cosmetic composition of claim 73 further contains a lung surfactant.

75. The cosmetic composition of claim 73, wherein the UG has a purity of about 75% to about 100%.

76. The cosmetic composition of claim 73, wherein the UG has a purity of about 90% to about 100%.

77. The cosmetic composition of claim 77 [sic], wherein the UG has a purity of at least 95%.

78. A blood supplement containing an amount effective to inhibit PLA2 of native or recombinant human uteroglobin (UG) or a derivative or fragment thereof, and a pharmaceutically acceptable carrier or diluent.

79. The composition of the blood complement of claim 78 further contains a lung surfactant.

80. The composition of the blood complement of claim 78, wherein the UG has a purity of about 75% to about 100%.

81. The composition of the blood complement of claim 78, wherein the UG has a purity of about 90% to about 100%.

82. The composition of the blood complement of the claim 78, wherein the UG has a purity of at least 95%.

83. A blood supplement composition containing an effective amount that binds to native or recombinant human uteroglobin fibronectin (UG) or a derivative or fragment thereof, and a pharmaceutically acceptable carrier or diluent.

84. The composition of the blood complement of claim 83 further contains a lung surfactant.

85. The composition of the blood complement of claim 83, wherein the UG has a purity of about 75% to about 100%.

86. The composition of the blood complement of claim 83, wherein the UG has a purity of about 90% to about 100%.

87. The composition of the blood complement of claim 83, wherein the UG has a purity of at least 95%.

88. An assay for quantifying uteroglobin-fibronectin complexes in a clinical sample, wherein a clinical specimen suspected to contain one or more uteroglobin-fibronectin complexes: (a) is contacted with an antigen-capturing agent, ( b) an agent for the detection of the antigen is addition to the sample, and (c) the presence of any complex bound to the capture agent is detected using a secondary antibody conjugated to a portion capable of being detected by the addition of a compound that reacts with the chemical moiety.

89. The assay of claim 88, wherein the agent for the capture of the antigen is a rabbit polyclonal, monospecific antibody, immobilized on an insoluble support.

90. The assay of claim 88, wherein the secondary antibody is an anti-IgG antibody conjugated to an enzyme and the compound that reacts with the chemical portion is a substrate for the enzyme that is converted to a chromogenic or fluorogenic compound with the reaction with the enzyme.

91. The assay of claim 90, wherein the enzyme is horseradish peroxidase.

92. The assay of claim 88, wherein the agent for the detection of the antigen is an antibody specific for fibronectin.

93. A method for the treatment or prevention of fibrillogenesis in a patient in need of such treatment, the method comprises the step of administering to a patient an effective amount for fibronectin binding to Native or recombinant UG or a fragment or derivative thereof.

94. The method of claim 93, wherein the UG has a purity of between about 75% to about 100%.

95. The method of claim 93, wherein the UG has a purity of between about 90% to about 100%.

96. The method of claim 93, wherein the UG has a purity of at least 95%.

97. The method of claim 93, where the UG is recombinant (rhUG) and has practically the same sequence as the native human uteroglobin.

98. The method of claim 97, wherein the effective amount of UG that binds fibronectin is 20 mg / kg to 500 mg / kg.

99. The method of claim 97, wherein the UG is administered in association with a lung surfactant.

100. The method of claim 99, wherein the lung surfactant is present in an amount of about 20% to about 80% by weight.

101. The method of claim 93, wherein the UG is administered by the endotracheal, ophthalmic, intravenous, intraperitoneal, intramuscular, liquid or oral routes.

102. The method of claim 93, wherein the UG It is administered as a liquid or semi-aerosol through an intratracheal tube.

103. The method of claim 93, wherein the fibrotic condition is pulmonary fibrosis.

104. The method of claim 93, wherein the fibrotic condition is renal fibrosis.

105. The method of claim 93, wherein the fibrotic condition is vascular fibrosis.

106. The method of claim 38, wherein the viral infection is CMV retinitis.

107. The method of claim 38, wherein the bacterial infection is pneumonia or cystitis.

108. The method of claim 38, wherein the parasitic infection is schistosomiasis.

109. The method of claim 38, wherein the fungal infection is vaginal candidiasis.