CN109846903A - A kind of set medicine of combination lung lavage treatment pulmonary fibrosis - Google Patents
A kind of set medicine of combination lung lavage treatment pulmonary fibrosis Download PDFInfo
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- CN109846903A CN109846903A CN201711240989.3A CN201711240989A CN109846903A CN 109846903 A CN109846903 A CN 109846903A CN 201711240989 A CN201711240989 A CN 201711240989A CN 109846903 A CN109846903 A CN 109846903A
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Abstract
The invention discloses a kind of set medicines of combination lung lavage treatment pulmonary fibrosis.The present invention protects a kind of set medicine for treating pulmonary fibrosis and/or alleviating pulmonary fibrosis patients pulmonary dysfunction, including mescenchymal stem cell and lung-douching fluid.The set medicine further includes ambroxol hydrochloride, frusemide, dexamethasone sodium phosphate, Ceftriaxone Sodium, aminophylline and Racanisodamine.The present invention is not found the problem in safety by zoopery and clinical research confirmation, clinical grade stem-cell therapy pulmonary fibrosis；And after stem-cell therapy degree of fibrosis lowered, tentative confirmation is effective.The present invention is for the treatment of pulmonary fibrosis and the specific important meaning of clinical application research of stem cell.
Description
Technical field
The present invention relates to a kind of set medicines of combination lung lavage treatment pulmonary fibrosis.
Background technique
Pulmonary fibrosis is the terminal stage of many pulmonary disease development, seriously endangers the health and postoperative rehabilitation of patient, still
Lack effective treatment method.Current main therapeutic strategy has: anti-inflammatory, anti-fibrosis and anti-oxidant etc..Conventional drug and
Nutrition treatment method cannot prevent the progression of fibrosis of lung, and whole-lung lavage (whole-lung lavage, WLL) is commonly called as " washing
Lung ", it can effectively be removed, and the dust of the protein-like substance, sucking that deposit in alveolar space and bronchiole, to gulp down pneumoconiosis bubble macrophage thin
Born of the same parents (pulmonary alveolar macrophage, PAM) and its cause inflammation generated, fibrogenic factor etc., play
Except the cause of disease, improvement respiratory function, alleviate the effect of symptom.But Massive whole-lung lavage can only alleviate the state of an illness, can not delay or
Reverse pulmonary fibrosis progress.
Mescenchymal stem cell have stronger self-renewal capacity, low immunogenicity, to the orientation chemotaxis of target tissue with
And the advantages that bystander effect, become the hot spot of cell therapy in recent years.Mescenchymal stem cell has multi-lineage potential
Adult stem cell has broad application prospects in tissue repair and gene therapy.
Summary of the invention
The object of the present invention is to provide a kind of set medicines of combination lung lavage treatment pulmonary fibrosis.
Present invention firstly provides a kind of for treating pulmonary fibrosis and/or alleviating pulmonary fibrosis patients pulmonary dysfunction
Cover medicine, including mescenchymal stem cell and lung-douching fluid.
The lung-douching fluid is physiological saline.
The mescenchymal stem cell is stored in physiological saline.
The dosage of the mescenchymal stem cell is 1 × 106A cell/kg weight.
The dosage of the lung-douching fluid is 1L-1.5L.
The set medicine further includes ambroxol hydrochloride, frusemide, dexamethasone sodium phosphate, Ceftriaxone Sodium, aminophylline and disappears
Revolve anisodamine.
The ambroxol hydrochloride is specially Ambroxol Hydrochloride Glucose Injection.The dosage of the ambroxol hydrochloride is
60mg.The method of application of the ambroxol hydrochloride is vein instillation.
The frusemide is specially furosemide injection.The dosage of the frusemide is 20mg.The application of the frusemide
Mode is intravenous injection.
The dexamethasone sodium phosphate is specially dexamethasone sodium phosphate injection.The dosage of the dexamethasone sodium phosphate
For 10mg.The method of application of the dexamethasone sodium phosphate is intravenous injection.
The Ceftriaxone Sodium is specially ceftriaxone sodium for injection.The dosage of the Ceftriaxone Sodium is 3.0g.It is described
The method of application of Ceftriaxone Sodium is vein instillation.
The aminophylline is specially aminophylline injection.The dosage of the aminophylline is 0.5g.The application of the aminophylline
Mode is vein instillation.
The Racanisodamine is specially 654-2 (anisodamine).The racemization Tangut Anisodus Radix
Base amount is 10mg.The method of application of the Racanisodamine is intravenous injection.
The application method of the set medicine are as follows: after patient's anesthesia, using lung-douching fluid (concretely physiological saline) lavation disease
Stove position (dosage concretely 1L-1.5L)；After lavation, irrigating solution is exhausted；To lesions position injection 5-10ml containing
State mescenchymal stem cell physiological saline (dosage of mescenchymal stem cell be 1 × 106A cell/kg weight).In surgical procedure
With the use of following drug: Ambroxol Hydrochloride Glucose Injection (ambroxol hydrochloride 60mg, vein instill), furosemide injection
(frusemide 20mg, intravenous injection), dexamethasone sodium phosphate injection (dexamethasone sodium phosphate 10mg, intravenous injection), injection
With Ceftriaxone Sodium (Ceftriaxone Sodium 3.0g, vein instill), aminophylline injection (aminophylline 0.5g, vein instill), 654-2
(anisodamine) (Racanisodamine 10mg, intravenous injection).
The present invention also protects the application of mescenchymal stem cell, for as follows (a1) or (a2):
(a1) product for treating pulmonary fibrosis is prepared；
(a2) pulmonary fibrosis is treated.
The present invention also protects the application of mescenchymal stem cell, for as follows (b1) or (b2):
(b1) product for alleviating pulmonary fibrosis patients pulmonary dysfunction is prepared；
(b2) alleviate pulmonary fibrosis patients pulmonary dysfunction.
The present invention also protects a kind of product, and active constituent is mescenchymal stem cell；The purposes of the product is treatment lung
Fibrosis.
The present invention also protects a kind of product, and active constituent is mescenchymal stem cell；The purposes of the product is to alleviate lung
Fibrosis patients' pulmonary dysfunction.
Any description above mescenchymal stem cell is clinical grade mescenchymal stem cell.
Any description above mescenchymal stem cell can be umbilical cord mesenchymal stem cells.The umbilical cord mesenchymal stem cells are behaved
Umbilical cord mesenchymal stem cells.Any description above mescenchymal stem cell can be to be passaged to for the 5th generation from what in vitro people's umbilical cord obtained
Mesenchymal stem cells.
The present invention by zoopery and clinical research confirmation, clinical grade stem-cell therapy pulmonary fibrosis in safety not
It finds the problem；And after stem-cell therapy degree of fibrosis lowered, tentative confirmation is effective.The present invention is for lung fiber
The treatment of change and the specific important meaning of clinical application research of stem cell.
Detailed description of the invention
Fig. 1 is canine leucocyte and cent lymphocytes inspection result in embodiment 1.
Fig. 2 is dog arterial blood gas analysis testing result in embodiment 1.
Fig. 3 is the content detection result of dog serum hydroxyproline and transforming growth factor in embodiment 1.
Fig. 4 is dog chest CT inspection result in embodiment 2.
Fig. 5 is canine leucocyte and cent lymphocytes testing result in embodiment 2.
Fig. 6 is dog arterial blood gas analysis testing result in embodiment 2.
Fig. 7 is the content detection result of dog serum hydroxyproline and transforming growth factor in embodiment 2.
Fig. 8 is dog serum c reactive protein and the horizontal testing result of IgG in embodiment 2.
Fig. 9 is 1 patient's blood routine, kidney function, myocardial enzymes testing result in embodiment 3.
Figure 10 is 1 Pulmonary Function testing result in embodiment 3.
Figure 11 is 1 Pulmonary Function testing result in embodiment 3.
Figure 12 is 1 Pulmonary Function testing result in embodiment 3.
Figure 13 is 1 patient chest CT testing result in embodiment 3.
Figure 14 is 1 patient chest CT testing result in embodiment 3.
Specific embodiment
Embodiment below facilitates a better understanding of the present invention, but does not limit the present invention.Experiment in following embodiments
Method is unless otherwise specified conventional method.Test material as used in the following examples is unless otherwise specified certainly
What routine biochemistry reagent shop was commercially available.Quantitative test in following embodiment is respectively provided with three repeated experiments, as a result makes even
Mean value.
Human umbilical cord mesenchymal stem cells in the present invention are by Chinese Academy of Sciences's heredity institute's stem cell bank from vitro people's umbilical cord
Middle to carry out separation and cultivate acquisition, identify through flow cytometry cell surface antigen: display CD14, CD34 and CD45 is yin
Property, CD29, CD44 and CD105 are the positive, are rendered as the feature of interstital stem cell.It is filled between people's umbilical cord used in the following embodiment
Matter stem cell is the human umbilical cord mesenchymal stem cells for being passaged to for the 5th generation.
The foundation of embodiment 1, pulmonary fibrosis animal model
One, experimental animal
Beasle dog: Beijing Marshall Biotechnology Co., Ltd, -12 monthly age of 7 monthly age, weight 7.0kg-12kg, male.It is qualified
Card number: SCXY (capital) 2002-0007.
Two, experimental drug
Yellow Jackets (Solution on Chemical Reagents in Shanghai Co., Ltd, Chinese Medicine group, lot number: WS20050411) use physiology salt
Water is configured to the pentobarbital sodium injection of 3% (mass percent), filtration sterilization, and the filled filter membrane of filter is bilayer, aperture
It is successively 0.45 μm and 0.22 μm, when filtering carries out in superclean bench.
Su-Mian-Xin mixture (Jilin Hua Mu animal health-care product Co., Ltd): by haloperidol, Baoding is peaceful, double hydrogen
Compound preparation made of the drugs such as Etorphine.
Three, the foundation of pulmonary fibrosis animal model
Beasle dog is grouped as follows at random:
Control group (8): by beasle dog fasting 12 hours (fasted overnight) before experiment.Using Su-Mian-Xin mixture
0.03ml/kg intramuscular injection, after dog is calm, the pentobarbital sodium injection 0.8ml/kg vein of 3% (mass percent) is infused
It penetrates, is given after anesthesia and pretend irradiation.
15Gy experimental group (8): by beasle dog fasting 12 hours (fasted overnight) before experiment.Using Su-Mian-Xin
Mixture 0.03ml/kg intramuscular injection, after dog is calm, the pentobarbital sodium injection 0.8ml/kg of 3% (mass percent) is quiet
Arteries and veins injection.Dog is fixed on experimental bench in dorsal position four limbs and is irradiated, radioactive source: 6MV x-ray linear accelerator, dosage rate
400cGy/min.Under radiotherapy simulation localization machine, determines and radiate wild (irradiation of half chest of right side), ource-skin Distance 100cm, exposure dose
15Gy irradiates 1 time.
Four, animal model metrics evaluation
The beasle dog of step 3 is detected for the 30th day, 90 days and 180 days as follows respectively before irradiation and after irradiation:
(1) leucocyte and cent lymphocytes
Beasle dog venous blood samples carry out white blood cell count(WBC) and lymphocyte hundred by blood-counter system (new bridge hospital)
Divide than checking.
As a result as shown in Figure 1.Medulla hematopoietic system is the sensitive organ of radiation, and the blood leukocytes of beasle dog are irradiated in 15Gy
Afterwards in 6-10 × 10 in 0-180 days9It is fluctuated within the scope of/L, belongs to the leucocyte normal range (NR) of beasle dog, lymphocyte percentage
It is not obvious than the variation before and after irradiation, the results showed that there is no systemic hematopoiesis functions for the local irradiation of 15Gy.
(2) arterial blood gas analysis
Femoral artery blood is extracted after phase point anaesthetizes beasle dog when each, with blood gas analyzer (U.S. GEM Premier 3000)
Lung function situation is observed in detection.
As a result as shown in Figure 2.The result shows that blood oxygen pressure is gradually reduced as time goes by, until 15Gy irradiates the 180th day
When, reduced about 50% (compared with before irradiation, p < 0.05), the variation of blood carbon dioxide partial pressure, HCO3- concentration and pH value compared with
It is small, it tends to be steady before and after irradiation.
(3) content of Serum oxyproline and transforming growth factor
Phase point measures the level of hydroxyproline in serum when each.When each phase point take hematometry serum transfer metaplasia long respectively because
Level (dog serum transforminggrowthfactor-α/β (TGF- α/TGF-β), c reactive protein and the IgG of son-α/β (TGF- α/TGF-β)
ELISA kit be purchased from Shanghai bioengineering Co., Ltd).
As a result as shown in Figure 3.The result shows that Serum oxyproline content gradually rises (Fig. 3 A) after irradiation, until after irradiation
At the 180th day, about 30ug/mL is had risen to, increases significant (p < 0.05) compared with before irradiation；Equally, seroconversion growth because
Sub- TGF-β after irradiation the 180th day when also obviously increase (p < 0.05), imply that degree of fibrosis aggravate (Fig. 3 C).Opposite,
The trend (Fig. 3 B) being gradually reduced is presented in serum TG F- α after irradiation.
The effectiveness and reliability evaluation test that embodiment 2, clinical grade mescenchymal stem cell treat pulmonary fibrosis
It is divided to the beasle dog after 8 15Gy experimental groups are irradiated 180 days in embodiment 1 to two groups of carry out pulmonary fibrosis treatments,
Specific step is as follows:
Physiological saline processing group (4): by fasting 12 hours (fasted overnight) before experiment, anesthesia (is infused using Su-Mian-Xin Ⅱ
Liquid mixture 0.03ml/kg intramuscular injection is penetrated, after dog is calm, the pentobarbital sodium injection 0.8ml/ of 3% (mass percent)
Kg intravenous injection) after, lavation is carried out using bottom right lung of the 100ml physiological saline to beasle dog.
HUCBMSCs transplantation group (4): by fasting 12 hours (fasted overnight) before experiment, anesthesia (is infused using Su-Mian-Xin Ⅱ
Liquid mixture 0.03ml/kg intramuscular injection is penetrated, after dog is calm, the pentobarbital sodium injection 0.8ml/ of 3% (mass percent)
Kg intravenous injection) after, the 5-10ml of lower lung injection to the right contains the physiological saline (stem cell population 1 of umbilical cord mesenchymal stem cells
×106A cell/Kg).
Respectively pulmonary fibrosis treatment before and treatment after being detected as follows within the 3rd day, the 90th day and the 180th day:
(1) chest CT examination
Experimental dog is under abundant anesthesia in southwestern hospital's row chest CT examination, the above specialist couple of You Liangwei attending physician
Chest CT is analyzed, including pulmonary shadow (including sheet shade, bar rope shadow, ground glass shadow, cavity shadow and consolidation shadow) and
Honeycomb shadow (including blood vessel bronchus beam is coarse, interlobuar septal thickening, subpleural line and net knit shadow).
As a result as shown in Figure 4.The result shows that the visible grid shadow of right lung CT result, interlobular septum are irregular before treating
It thickens, tiny blood wall thickening is obvious in leaflet, and the lower partially visible honeycomb shade of pleura pulmonalis, illustrates radiation fibrosis of lung in the right side
Modeling is set up.After mescenchymal stem cell is treated, the extension of honeycomb shade at any time gradually subtracts right lung CT as the result is shown
It is few, imply that degree of injury gradually mitigates；And after physiological saline is handled, obviously expand in the 30th day honeycomb shade, but
It is to extend as time go on, shaded area is also being gradually reduced, and illustrates that degree of injury is equally mitigating, but still fills between overweighting
Matter stem cell transplantation group.
(2) leucocyte and cent lymphocytes
Detection method is the same as embodiment 1.
As a result as shown in Figure 5.The result shows that either physiological saline processing or mesenchymal stem cell transplantation, it is unknown
Develop and rings peripheral white blood cells and cent lymphocytes, in 180 days after treatment, peripheral white blood cells and lymphocyte hundred
Divide and is fluctuated than being in normal range (NR).
(3) arterial blood gas analysis
Detection method is the same as embodiment 1.
As a result as shown in Figure 6.
The result shows that blood oxygen pressure gradually gos up as time goes by after mesenchymal stem cell transplantation, with physiological saline group
Between significant difference (p < 0.05), show that mesenchymal stem cell transplantation can improve lung function；And blood carbon dioxide partial pressure, HCO3- are dense
The variation of degree and pH value is smaller, no significant difference (p > 0.05) between physiological saline group.
(4) content of Serum oxyproline and transforming growth factor
Detection method is the same as embodiment 1.
As a result as shown in Figure 7.The result shows that becoming for decline is presented in Serum oxyproline content after mesenchymal stem cell transplantation
Gesture (Fig. 7 A) reduces significant (p < 0.05) compared with physiological saline group；Equally, seroconversion growth factor TGF-β is in mesenchyma
It is significantly reduced (p < 0.05) after stem cell transplantation, implys that degree of fibrosis mitigates (Fig. 7 C).Opposite, serum TG F- α exists
The trend (Fig. 7 B) gradually increased is presented after mesenchymal stem cell transplantation.
(5) serum C-reactive protein and IgG are horizontal
It is operated, is examined in strict accordance with kit specification by ELISA kit (Shanghai bioengineering Co., Ltd)
The level of phase point serum C-reactive protein and IgG when surveying each.
As a result as shown in Figure 8.The result shows that the immune response water of body can not be improved after mesenchymal stem cell transplantation
It is flat, serum C-reactive protein and IgG the level equal no significant difference (p > 0.05) compared with before physiological saline group and itself processing.
Embodiment 3, clinical grade umbilical cord blood mesenchymal stem cells treat the clinical research of pulmonary fibrosis
One, research object
Research object: 8 clinical diagnosises are the patient of pulmonary fibrosis.
Two, surgical procedure
After chest CT positioning, patient's horizontal position uses the slow lavation lesions position of physiological saline under local anaesthesia, every time 50 milli
It rises, amounts to 1.0-1.5 liter.After lavation, irrigating solution is exhausted as far as possible.It is 92 or more and life to patient's transcutaneous oxygen saturation
After life sign is steady, the 5-10ml physiological saline for containing clinical grade human umbilical cord mesenchymal stem cells is injected into disease through Bronchofiberscope again
(usage amount of human umbilical cord mesenchymal stem cells is 1 × 10 for stove position6A cell/Kg).It is used cooperatively in surgical procedure as put in poison
Object: Ambroxol Hydrochloride Glucose Injection (ambroxol hydrochloride 60mg, vein instill), furosemide injection (frusemide 20mg, it is quiet
Arteries and veins is injected), dexamethasone sodium phosphate injection (dexamethasone sodium phosphate 10mg, intravenous injection), ceftriaxone sodium for injection (head
Spore Qusong sodium 3.0g, vein instill), aminophylline injection (aminophylline 0.5g, vein instill), 654-2 (hydrochloric acid racemic Tangut Anisodus Radix
Alkali injection) (Racanisodamine 10mg, intravenous injection).
Three, Prognosis scoveillance
1, Follow-up After has no special bad complication, and original symptom is showed without exacerbation.Completion follow-up patient, which has no, to be appointed
He Xinfa symptom feels shortness of breath symptom and is improved.Most SGRQ scoring gaps are greater than 4 points.
2, patient is 3 days postoperative, March and follow-up in June blood routine, kidney function, myocardial enzymes.1 patient is 3 days postoperative, March and 6
Moon follow-up blood routine, kidney function, each Indexs measure result of myocardial enzymes are as shown in figure 9, N indicates each index range of normal value, as a result
Show each index in the normal range.Other patients are 3 days postoperative, March and each index of blood routine in June, kidney function, myocardial enzymes
In the normal range.
3, patient is preoperative, postoperative March and follow-up in June lung function.Wherein, pulmonary function detection result such as Figure 10 of 1 patient
Shown, the patient is preoperative, the lung function ventilation index actual value in postoperative March and June does not significantly reduce.The lung function of 1 patient
Can testing result it is as shown in figure 11, the patient is preoperative, the lung function ventilation index actual value in postoperative March and June does not obviously subtract
It is few.The pulmonary function detection result of 1 patient is as shown in figure 12, and the lung function of the patient changes without particular law, but with preoperative ratio
Relatively aggravated without obvious.Other patients are preoperative and postoperative March, lung function in June are without significant change.
4, follow-up of patients's chest CT.The CT result of 1 patient is as shown in figure 13.It is from left to right preoperative, postoperative in Figure 13
January and June.No. 1 patient's arrow meaning is diseased region, which is patient with breast cancer, and radiation fibrosis of lung degree is low, warp
After treatment, fiber foci disappearance is visually observed.The CT result of 1 patient is as shown in figure 14.In Figure 14, from left to right for it is preoperative,
Postoperative January and June, patient's arrow meaning are diseased region, and visually observing lesion has absorption.Other patient's degree of fibrosiss are equal
Lowered.
In summary zoopery and clinical research confirmation, clinical grade stem-cell therapy pulmonary fibrosis are not sent out in safety
Existing problem；And after stem-cell therapy degree of fibrosis lowered, tentative confirmation is effective.
Claims (10)
1. a kind of set medicine for treating pulmonary fibrosis and/or alleviating pulmonary fibrosis patients pulmonary dysfunction, including mesenchyma are dry
Cell and lung-douching fluid.
2. set medicine as described in claim 1, it is characterised in that: the lung-douching fluid is physiological saline.
3. set medicine as claimed in claim 1 or 2, it is characterised in that: the mescenchymal stem cell is stored in physiological saline.
4. the set medicine as described in claims 1 to 3 is any, it is characterised in that: the dosage of the mescenchymal stem cell is 1 × 106It is a
Cell/kg weight.
5. the set medicine as described in Claims 1-4 is any, it is characterised in that: the set medicine further includes ambroxol hydrochloride, furan plug
Rice, dexamethasone sodium phosphate, Ceftriaxone Sodium, aminophylline and Racanisodamine.
6. the application of mescenchymal stem cell, for as follows (a1) or (a2):
(a1) product for treating pulmonary fibrosis is prepared；
(a2) pulmonary fibrosis is treated.
7. the application of mescenchymal stem cell, for as follows (b1) or (b2):
(b1) product for alleviating pulmonary fibrosis patients pulmonary dysfunction is prepared；
(b2) alleviate pulmonary fibrosis patients pulmonary dysfunction.
8. a kind of product, active constituent is mescenchymal stem cell；The purposes of the product is treatment pulmonary fibrosis.
9. a kind of product, active constituent is mescenchymal stem cell；The purposes of the product is to alleviate pulmonary fibrosis patients lung function
It can obstacle.
10. set medicine as claimed in claim 1 to 5, or, application described in claim 6 or 7, or, claim 8 or 9
The product, it is characterised in that: the mescenchymal stem cell is umbilical cord mesenchymal stem cells.
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| CN113018309A (en) * | 2021-02-09 | 2021-06-25 | 佛山市第二人民医院（佛山市便民医院） | Lavage fluid and preparation method and application thereof |
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