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 ::  Abstract
 ::  Introduction
 ::  Methods
 ::  Results
 ::  Discussion
 ::  References
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PAPERS
Year : 1994  |  Volume : 40  |  Issue : 2  |  Page : 65-7

Modulation of Kupffer cell activity by Tinospora cordifolia in liver damage.


Dept of Pharmocology, Seth GS Medical College, Parel, Bombay.

Correspondence Address:
D S Nagarkatti
Dept of Pharmocology, Seth GS Medical College, Parel, Bombay.

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Source of Support: None, Conflict of Interest: None


PMID: 0008737554

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 :: Abstract 

Kupffer cells are major determinants of outcome of liver injury. Their activity was therefore studied in a model of chronic liver disease. The effect of Tinospora cordifolia, an indigenous agent with proven hepatoprotective activity, was evaluated on Kupffer cell function, using carbon clearance test as a parameter. Rats were divided into two major groups. In Gp I which served as normal control t1/2 of carbon was 9.48 +/- 4.14 min. GpII received horse-serum in a dose of 0.5 ml/100 gm b.w. i.p. for a period of 12 weeks and was divided into three sub-groups. In Gp IIA at the end of 12 weeks half-life of carbon was found to be significantly increased to 19.86 +/- 7.95 min (p < 0.01). Indicating suppressed Kupffer cell function in chronic liver damage. In Gp IIB treated with vehicle for 4 more weeks there was significant prolongation of half-life to 38.32 +/- 10.61 min (p < 0.01), indicating perpetuation of damage in absence of damaging agent. Whereas in Gp IIc, treated with Tinospora cordifolia t 1/2 was decreased to 14.24 7.74 min (p < .01), as compared to vehicle control indicating a significant improvement in Kupffer cell function and a trend towards normalization.


Keywords: Analysis of Variance, Animal, Carbon, pharmacokinetics,Disease Models, Animal, Female, Kupffer Cells, physiology,Liver Failure, metabolism,physiopathology,therapy,Male, Metabolic Clearance Rate, Plants, Medicinal, Rats,


How to cite this article:
Nagarkatti D S, Rege N N, Desai N K, Dahanukar S A. Modulation of Kupffer cell activity by Tinospora cordifolia in liver damage. J Postgrad Med 1994;40:65

How to cite this URL:
Nagarkatti D S, Rege N N, Desai N K, Dahanukar S A. Modulation of Kupffer cell activity by Tinospora cordifolia in liver damage. J Postgrad Med [serial online] 1994 [cited 2020 Apr 1];40:65. Available from: http://www.jpgmonline.com/text.asp?1994/40/2/65/562





  ::   Introduction Top


Chronic liver damage in humans and in the experimental animal is characterized by deposition of fibrous tissue in liver. Prognosis of chronic liver damage goes on worsening with increase in fibrous tissue content of liver[1]. With the advent of refined biochemical and ultra-structural methodologies, it has been proved that deposition of fibrous tissue in chronic liver damage is a result of disturbance in the regulatory network between various liver Cells[2]. Of these cells, Kupffer cells are considered as major determinant of outcome of liver injury[3]. Under normal circumstances, these cells keep an inhibitory control on the Ito cells, the precursor fibroblasts of the liver. A suppressed activity of Kupffer cells therefore leads to proliferation and activation of Ito Cells[4] resulting in fibrous tissue deposition[5]. Hence, it appears that therapy directed at activating Kupffer cells will have anti-fibrotic potential.

In our laboratory, Tinospora cordifolia, a plant belonging to the family Menispermiacae, has been shown to decrease the fibrosis in animal models of reversible and irreversible liver injuries induced by carbon tetrachloride[6] as well as heterologous serum[7]. This antif ibrotic effect was found to preserve the liver architecture as judged by histopathology and hydroxyproline content of liver. This plant has also been shown to increase phagocytic and killing activity of macrophages, tissue elements of RES against various bacteria and fungi induced[8],[9].

Since the drug has shown anti-fibrotic effect and also found to be an activator of macrophages, it was of interest to determine whether it can modulate the activity of Kupffer cells, tissue macrophages of liver, in chronic liver damage. Hence, the following study was undertaken.


  ::   Methods Top


A model of fibrosis simulating post-hepatitis cirrhosis was set up. The activity of Kupffer cells in rat fibrotic liver was determined and compared with that of normal. The effect of Tinospora cordifolia therapy on fibrosis was evaluated with respect to activity of Kupffer cells. Twenty-four rats of either sex weighing between 200-500 gms were given injections of horseserum in the dose of 0.5m1/kg b.w. i.p. twice a week for 12 weeks[10] (Gp II). A group of 8 normal rats matching in age and sex was maintained throughout this period without any treatment (GpI). At the end of 12 weeks, Kupffer cell activity, of 8 out of 24 rats (GpIIA) was measured by determining carbon clearance and compared with that of normal (Gpl).

The remaining 16 rats were divided into two groups after 12 weeks of horse-serum injections. One group received vehicle (GpIIB) and the other was treated with Tinospora cardifolia (GpIIc), both for a period of 4 weeks. Powdered stem of Tinospora cordiofolia was mixed with water and a decoction was prepared. The dose selected was 100 mg of powdered stem/kg b.w. orally once a day. At the end of 4 weeks activity of Kupffer cells was assessed.

To study the Kupffer cell activity in the above mentioned study, carbon clearance method as described by Heller (1958)[11] was used. Initially a series of dilutions of colloidal carbon (donated by Camlin Ink) were prepared. The optical density (O.D.) of these dilutions was measured using a spectrophotometer at a wavelength of 650nm with a 1cm path length. This procedure was repeated by adding 50 ?l of blood (collected from rats) to the various dilutions of carbon and processing the samples as mentioned below. After obtaining the optical densities of these dilutions a standard curve of concentration of carbon vs optical density was constructed (Fig 1).

Blood samples were obtained from retro-orbital sinus of all the rats under test with 50?l capillaries (0 min), following which each rat was injected colloidal carbon in a dose of 8mg/100gm b.w.i.v. through femoral vein. Serial blood samples were collected at intervals of 5, 10, 15, 20,25 min., from retro-orbital sinuses with 50?l capillaries and lysed in 4ml of sodium carbonate (0.1%) solution and O.D. of these solutions were determined. These O.D. readings were used in conjunction with standard curves to calculate the amount of carbon in the blood. The concentration -time curves were plotted from which, half-life of carbon particles was calculated using least square method. This half-life of carbon particles indicates the activity of Kupffer cells.

To compare activity & Kupffer cells from different groups GpI, Gp IIA, Gp IIB, Gp IIC statistical analysis was performed by use of analysis of variance (ANOVA). If significant differences between groups were identified by ANOVA, specific comparisons between individual group were accomplished by Student's unpaired test. Results are presented as mean + S.D.


  ::   Results Top


The mean optical density - concentration curve for carbon particles is depicted in [Figure:1].

The sensitivity of this test is 2.5 ng/ml. The coefficient of correlation was 0.99. The coefficients of variation for the different concentrations of carbon are as follows: 2.81%, 2.85%, 3.55%, 3.63, 2.99%, 2.21%, 2.34% for 5, 7.5, 10, 12.5, 15, 20, 22.5, 25 respectively.

The results are shown in [Table - 1].



The half-life of four groups differed significantly by ANOVA (p < 0.01). The carbon clearance in normal rats (GpI) was 9.48 + 4.14 min. In rats receiving horse-serum for 12 weeks (GpIIA) the half-life was significantly prolonged to 19.96 + 7.65 min (p < 0.01). In the rats who were administered vehicle therapy (GpIIB) forturther period of 4 weeks following horse-serum injections the half-life was found to be prolonged further to 38.32 + 10.61 min (p < 0.01 as compared to GpIIA), whereas in rats treated with Tinospora (GpIIC) for an equivalent period the half-life of carbon was found to be 14.24 + 7.74 min (p < 0.01 as compared to GpIIB).


  ::   Discussion Top


Present study was carried out to evaluate effect of Tinospora cordifolia on Kupffer cell activity in a model of chronic liver damage. Such model of fibrosis of liver was induced in rats by repeatedly injecting horseserum. Antibodies developed against this heterologous serum combine with antigen andensuesan inflammatory process. The sequelae of such inflammation is fibrosis[12].

In the present study following horse-serum injection the half-life of carbon was found to be significantly prolonging (GpII A), which in turn indicates suppressed Kupffer cell activity in these animals. Depressed activity of Kupffer cells in chronic liver damage has also been reported by Noda et al[13].

It was observed in the present study that even if the damaging agent is withdrawn (and rats were left untreated for next 4 wks i.e. GpII B), the activity of Kupffer cells did not improve; in fact clearance of carbon was found to be significantly reduced (as judged 3. by the prolonged half-life) as compared to GpII A. Thus further suppression of Kupffer cell activity was noticed in GpIIB, which received no therapy.

On the other hand, in GpIIC the half-life of carbon clearance was found significantly less than the untreated GpIIC and comparable to control group. This indicates that Tinospora therapy has prevented deterioration of Kupffer cell activity. This appears to be due to its macrophage activating property, which has opposed any suppressant influences on Kupffer cells.

Currently a lot of research in the field of hopatology has been directed to evaluate the relation between Kupffer cell activity and development of hepatic fibrosis. It has been proved that suppression of Kupffer cells deprives the hepatocytes of cytoprotective effect[14], releases inhibitory control over Ito Cells[4], disrupts detoxification of endotoxins[15] and decreases capacity to secrete collagenase[16].

In our previous study[7] we have confirmed the deposition of fibrous tissue both by histopathological examination and by estimation of liver hydroxyproline[7]. On discontinuation of horse serum, perpetuation of fibrotic process was observed in damaged liver[7]. Such a phenomenon has been reported previously[12]. Therapy with Tinospora cordifolla has been shown to prevent the fibrous tissue deposition in the post insult period[7].

Findings of our present study raise a possibility that this anti-fibrotic effect of Tinospora cordifolia is mediated through activation of Kupffer cells.

 
 :: References Top

1. Hahn EG, Schuppan D. Collagen metabolism in liver disease. In Bianchi L. Greok W, Landmann L, Sickinger K, Staider OA, editors. Liver in Metabolic Disease, Proceedings of 35th galk symposium Base1. MTP Press Ltd; 1983, pp 309-23.  Back to cited text no. 1    
2.Popper H. Regulatory modulation in hepatology. Hepatology 1987; 7:86-90.   Back to cited text no. 2    
3.Bjorneboe M, Prytz H. The mononuclear phagocytic functions of liver. In: Fergusson A, Maclween RNM, editors. Immunobiological Aspects of the Liver and GI Tract. England: MTP Press Ltd; 1976, pp 250-288.  Back to cited text no. 3    
4.Rojkind M, Valadez G, Rosas M. Cell -Cell homeostasis in normal and fibrotic rat liver Hepatology 1983; 3:851-854.  Back to cited text no. 4    
5.Ballardini G, Esposti SD, Bianchi FB. Correlation between Ito cells and fibrogenesis in an experimental model of hepatic fibrosis. A sequential stereological study. Liver 1983; 3:58-63.  Back to cited text no. 5    
6.Rege NN, Dahanukar SA, Karandikar SM. Hepatoprotective effect of Tinospora corditolia against carbon tetrachloride induced liver damage. Indian Drugs 1984; 21:544-555.  Back to cited text no. 6    
7.Malagi S. Modulation of liver injury by plant products. MSc thesis 1989.  Back to cited text no. 7    
8.Rege NN, Dahanukar SA. Quantitation of microbicidal activity of mononuclear phagocytes: an in vitro technique. J Postgraduate Med 1993; 39:22-25.  Back to cited text no. 8    
9.Thatte UM, Dahanukar SA. Immunotherapeutic modification of experimental infections by Indian medicinal plants. Phytother Res 1989; 3:43  Back to cited text no. 9    
10.Paronetto F, Popper H. Chronic liver injury induced by immunologic reactions. Am J Pathol 1966; 49:1087-1101.  Back to cited text no. 10    
11.Helier JH. Measurement of function of RES Ann New York Acad Sci 1958; 73:212-214.  Back to cited text no. 11    
12.Thomas HC. Immonological aspects of hepatic fibrosis Br J Gastroenterol 1980; 12:33- 6.13.  Back to cited text no. 12    
13.Noda T, Mimura H, Orita K. Assessment of Kupffer cell function in rats with chronic liver injury caused by CCI. Hepatogastroenterology 1990; 37:319-323.  Back to cited text no. 13    
14.Decker K. Eicosanoids, signal molecules of liver cells. Semin Liver Dis 1985; 5:185-190.  Back to cited text no. 14    
15.Toth CA, Thomas P. Liver endocytosis and Kupffer cells. Hepatology 1992; 16:255-266.  Back to cited text no. 15    
16.Montfort I, Perez-tamayo R. Collagenase in experimental carbon tetrachloride cirrhosis of the liver. Am J Pathol 1978; 92:411-420.   Back to cited text no. 16    


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Online since 12th February '04
2004 - Journal of Postgraduate Medicine
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