JPS61290961A - Artificial liver apparatus - Google Patents

Abstract

(57) [Summary] This bulletin contains application data before electronic filing, so abstract data is not recorded.

Description

[Detailed description of the invention] [Industrial application field] The present invention relates to an artificial liver device made of an adsorbent or the like.

More specifically, the present invention relates to an artificial liver device for treating liver failure, which is comprised of a cation exchanger as an adsorbent and means for interfering with the cation exchanger.

[Prior Art] The liver is an important organ that performs functions such as metabolism and synthesis of various substances, and detoxification and conjugation of intermediate metabolites and drugs. Therefore, liver disease has a serious impact on the maintenance of life.

In particular, acute liver failure is feared because of its high mortality rate.

Various artificial liver devices are being devised for such cases.

However, none of the current artificial liver devices has been able to demonstrate definitive effectiveness. For example, artificial liver devices that utilize the metabolic ability of living liver cells of xenobiotic animals are being developed, but it is difficult to say that the capabilities of such devices have been fully utilized. Today, the understanding of the substances that should be supplied to the patient from hepatocytes or the substances that should be metabolized by hepatocytes has not progressed sufficiently, making it difficult to appropriately evaluate or optimize the design of such devices. It is.

Furthermore, adsorbents, particularly activated carbon or polystyrene-based anion exchange resins, are already in clinical use as artificial liver devices.

Various substances are already known to accumulate in the body during acute liver failure. Among these substances, free fatty acids, phenols, mercaptans, etc. can be removed by activated carbon.
Furthermore, bilirubin, bile acids, and the like are each well adsorbed by anion exchange resins. However, although these adsorptive artificial livers are partially effective, they have not been able to significantly improve the survival rate due to insufficient removal of toxicity.

[Problem that the invention seeks to solve]

An object of the present invention is to provide an excellent artificial liver device that can remove toxic components that could not be removed using conventional adsorption-type artificial livers.

[Means for solving problems]

The above object is achieved by the present invention as described below.

That is, the artificial liver device of the present invention is characterized by comprising the following parts.

(a) a means for lowering the pH of blood; (b) an adsorption device comprising a hydrophilic weakly acidic cation exchanger as an adsorbent; (c) a means for restoring the pH of blood; and (d) a means for reducing blood pH. The blood is treated with the pH lowering means, the adsorption device and the pH lowering means.
A liquid delivery mechanism for returning the liquid to the patient via the recovery means in this order.

Next, each part constituting the present invention will be explained.

a) pH lowering means: Any means can be used as long as the pH can be lowered, such as adding a weak acid or dialysis with a dialysate to lower the pH.
H may be lowered, but λpH is preferably lowered to 6.5 or less, particularly preferably 6.0 to 6.3. pH
It is even more preferable to lower the ionic strength of the blood at the same time.
It is preferable to lower the ionic strength to 0.1-0.15.

The most preferable dialysis device as a pH lowering means will be specifically explained below.

A dialysis machine is any device that can accept blood from a patient, lower its pH, and then pump it out continuously.

The device can be selected from those used in dialysis therapy as artificial kidneys. This can be achieved by receiving and flowing blood from the patient on one side of the dialysis membrane and flowing dialysate, which has a lower pH than the patient's blood, on the other side. The specific structure of the dialysis machine is, for example,
There are laminated flat plate types, hollow fiber types, etc., and the details are described in "Igaku no Ayumi, Vol. 105, No. 5, 497 (1978)".

The dialysate may be any fluid that can lower the p)I and ionic strength of the patient's blood. For example, it may be a phosphate buffer plus sodium chloride. However, from the viewpoint of minimizing the burden on the patient, a commercially available dialysate used in artificial kidneys to which water and acetic acid have been added is preferably used.

The specific prescription of the dialysate is described in the catalog of "Kindery Fluid AP-1" by Fuso Pharmaceutical Co., Ltd.

b) Adsorption device: The adsorption device receives patient blood at a reduced pH and
This is a device that can adsorb positively charged proteins. Since this device is used for extracorporeal circulation, the material for the inner wall of the device and the adsorbent are selected from materials that do not denature or otherwise damage plasma proteins and are highly compatible with blood.

That is, the material for the inner wall of the device is selected from existing antithrombotic materials, and the adsorbent is selected from hydrophilic and weakly acidic cation exchangers.

As the hydrophilic weakly acidic cation exchanger, insoluble poly#N derivatives are preferable, and specifically, crosslinked dextran derivatives such as carboxymethyl-Sephadex (Pharmacia), crosslinked carboxymethyl-Sepharose (Pharmacia), etc. Examples include agarose derivatives, carboxymethyl-Biogel A (Bio-Rad), and the like.

Although any of these can be used, carboxymethyl-Sepharose is preferred from the following two points of view. One is that it has a large exchange capacity for proteins, and the other is that it has mechanical strength so as not to cause clogging, and does not release particulates.

c) pH recovery means: The pH of the blood lowered by the means of a) is reduced to the pH of the patient's blood.
Although there are no particular limitations as long as it is a means for returning the blood to a certain point, a method using dialysis is most preferred, and a dialysis device will be specifically described below.

The dialysis machine is any device that can receive blood from an adsorption device, restore its pf (and ionic strength) to that of the original patient, and then pump it out continuously. Just like the dialysis device in a), the artificial kidney can be selected from those used in dialysis therapy.

One side of the dialysate receives and flows the blood from the adsorption device, and the other side receives the dialysate (which has a higher pH than this blood).
If the ionic strength is also lowered in the dialysis in a), the purpose can be achieved by flowing a dialysate with a higher ionic strength).

The dialysate may be any fluid capable of restoring the pH (and ionic strength) of the blood from the adsorption device to those of the original patient, respectively. for example,
It can be a phosphate buffer plus sodium chloride. However, from the viewpoint of minimizing the burden on the patient, commercially available dialysis fluids used in artificial kidneys are preferably used. The specific formulation of the commercially available dialysate is described in the catalog of "Kindery Fluid AF-1" by Fuso Pharmaceutical Co., Ltd.

d) Liquid feeding mechanism: The liquid feeding mechanism consists of piping and a pump that are connected so that blood from the patient sequentially and continuously flows through each of the above sections and finally returns to the patient.

Piping should have wetted surfaces of materials that are non-hazardous to blood. Specifically, there are vinyl chloride blood circuits used in artificial kidneys.

The pump can also be any used in artificial kidneys. Further, it is more preferable to attach a pH meter and an electrically conductive needle between each of the above-mentioned parts in order to continuously monitor the pH and ionic strength. Hereinafter, the artificial liver device according to the present invention will be described in more detail with reference to Examples.

Example 1 (1) Collection of liver failure model rat serum and evaluation of its cytotoxicity Galactosamine (1.25 g/kg body weight) was added to male Wis.
Tar rats were injected into the peritoneal cavity, blood was collected 24 hours later, and serum was prepared. Cytotoxicity evaluation was performed as follows. first,
A serial dilution series of 2-fold, 4-fold, 8-fold, 16-fold, etc. was prepared for the sample rat serum in each column of a 96-well plate using MEM medium supplemented with 5% calf serum as a diluent. For 100 μl of diluted rat serum in each well, add 100 μl of mouse L cells suspended in the above medium at a ratio of 104 cells/m1! In addition, the cells were cultured for about 1 week at 37°C in a carbon dioxide gas incubator.

The cell layer adhering and proliferating at the bottom of each well was stained with formalin-crystal violet according to a conventional method, and the absorbance of each well was colorimetrically determined using multiscan. The dilution concentration (%) of the sample serum exhibiting 50% absorbance of the control well containing only the medium was calculated and used as an index of toxicity.

Incidentally, even serum obtained from normal rats shows non-specific growth inhibition on cells in a high concentration range.

In normal rat serum, the serum concentration that reduces the number of L cells by 50% is 3-4%. Therefore, in the following experiment, if the concentration that increases the cell number to 50% increases to this extent, it can be considered that the above-mentioned factors have been completely adsorbed.

(2) Effects of dialysate pH and ionic strength on the adsorption of cytotoxic substances in rat serum to the adsorbent How far should the pH and ionic strength be lowered in order to completely adsorb cytotoxic factors? investigated.

Various dialysis solutions shown in Table 1 were prepared based on 20mM phosphate buffer with addition of sodium chloride. Add 1/100 volume of the dialysate of galactosamine-administered rat serum to a cellulose tube (
Union Carbide) and dialyzed overnight against these dialysates.

33 on dry weight basis for 1 ml of serum before and after dialysis
．． Carboxymethyl Sepharose CL at a rate of 3 mg
-68 (Pharmacia) was added and kept at 37°C for 2 hours. Evaluate cytotoxicity before and after dialysis and before and after adsorption,
The results are summarized in Table 1 along with biochemical data.

As can be seen from Table 1, dialysis with a dialysate having a pH of 6.0 and an ionic strength of 0.100 was optimal for complete adsorption. At this time, the pH of the serum was 6.24, the ionic strength was 0.145, and the serum concentration at 50% of the cells after adsorption was 3.86%. That is, the level was that of normal serum.

(3) Artificial liver device model experiment Based on the above data, a circuit simulating extracorporeal circulation was constructed to flow rat serum, and it was examined whether cytotoxicity in the serum could be removed.

The model circuit is shown in the figure. Serum is pumped up from a galactosamine-administered rat serum storage tank 2 by a pump 1 and sent to a first dialysis device (p11 lowering means) 4 via a blood circuit 3. The serum is further led in series to an adsorption device 5 and a second dialysis device (pH recovery means) 6, and finally returns to 2. ? ,8,
9.10 is a sampling boat.

The first and second dialysis devices used in this experiment were:
Toshi PMMA hollow fiber module B-1 (area 1.1
or). The first dialysis machine uses Kinderly fluid AF-1 (manufactured by Fuso Pharmaceutical) as an external fluid instead of the specified prescription.
Liquid: B liquid: Dilution water = 1: 1.26: 55.
The ionic strength is 0.101), and p
) I was set to 6.2 by adding acetic acid and perfused with 50011 Il/llin.

For the second dialysis machine, prepare Kinderly fluid meter No. 1 as an external fluid according to the specified prescription.Ionic strength 0, 165), p
Same as when H was adjusted to 7.8 with sodium bicarbonate <
Perfusion was performed at 500ml/win. As an adsorption device, a silicon-treated glass column (diameter 5 cm, length 10
cm) was filled with carboxymethyl-Sepharose CL-6B with nylon mesh sandwiched between the top and bottom. A circuit was formed using a vinyl chloride tube, and galactosamine-administered rat serum was allowed to flow at a rate of 30 III/min.

Once steady state was reached, samples were taken from each boat for analysis and evaluation. The results are shown in Table 2.

Table 2 Cytotoxicity of Serum in the Circuit The results above show that cytotoxicity found in patient or animal serum can be continuously removed without major changes in the final pH and ionic strength of the serum. That is clear.

[Brief explanation of the drawing]

The figure is a model circuit diagram of an artificial liver device according to the present invention. 1-・-・・Pump 2-・−・−Galactosamine-administered rat serum reservoir 3・-
・-Blood circuit 4--First dialysis device (pH lowering means) 5--11111 Adsorption device

Claims (1)

[Scope of Claims] (a) means for lowering the pH of blood, (b) an adsorption device using a hydrophilic weakly acidic cation exchanger as an adsorbent, (c) means for restoring the pH of blood, and (d) The blood from the patient is transferred to the pH lowering means, the adsorption device and the pH
An artificial liver device comprising: a feeding mechanism for returning fluid to a patient via a recovery means in this order;