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Selective intestinal decontamination with trimethoprim-sulfamethoxazole in an experimental model of cirrhosis in rats
HEPATOLOGY Vol. 22, No. 4, Pt. 2, 1995
1265
423A
AASLD ABSTRACTS
HEPATITIS B VIRUS VARIANTS WITH DEFECTS IN THE PRECORE REGION ARE INFECTIOUS IN VITRO. 1phiLinne Gring~, 2,3Shu-Ping Tong. 2Christian Treoo. and 1Christiane Guguen-Guillouzo
1266
1 Unit6 de recherche h6patologique U 49, Institut national de la sant6 et de la recherche m6dicaie, H6pital de PontchaiUou, avenue Henri le Guillou, 35053 Rennes Cedex, 2 Unit6 de recherche sur les h6patites, le SIDA et les r6trovirus humains U 271, Institut national de la sant6 et de la recherche m6dieale, 151, cours Albert Thomas 69424 Lyon Cedex 03, France, 3 Present address: Molecular Hepatology Lab, Massachusets General Hospital Cancer Center, Charlestown, MA 02129.
MILD OR AGGRESSIVE CHRONIC ACTIVE HEPATITIS (CAll) DUE TO HEPATITIS C VIRUS (HCV) ARE ASSOCIATED WITH DIFFERENT COMBINATIONS OF HCV GENOTYPES. ~,~derilflfi.I.Ferretti.A.Ferrari.P .Trande.R.Cosenza.F.Callea*.F .Maaenti.E.Villa. Gastroenterology, Univ.Modena *Spedali Civili,Brescia,Italy. CAH due to HCV infection is o~ten characterized histologically either by very mild inflammatory condition or by a very aggressive pioture, without intermediate situations. As the clinically these patients do not differ significantly, we investigated the details of HCV infection (levels of viremia and genotypes). We studied 52 consecutive patients who had the first recogaitiun of altered ALTs in the previous year, all with histological diagnosis of CAll: 24 had a very mild or borderline CAll (Kundell's score <5) and 27 a n aggressive hepatitis (Knodell's score >9). Viremia was investigated by RT/PCR with endpoint dilutions, HCV genotype by nested PCR according to Okamoto. Biochemical data were not significantly different in the two groups; mean age in mild CAH was significantly younger in comparison with the aggressive CAH (32+10 vs 46_+9). Viremia was not significantly different (mild CAH vs aggressive CAll: 106+102 vs 105+102). HCV genotypes were as follows:
HBV variants defective for e antigen secretion have been frequendy evidenced in blood samples. These variants show several types of mutations, leading to creation of a stop codon or elimination of the initiation codon in the precore region. To determine whether such mutants were still capable of supporting a complete replication cycle we have used an in vitro test system. Mutant viral DNAs bearing a representative panel of mutations in the precore region were transfected in HepG2 cells. The viral particles secreted in the supernatant were analyzed and further tested for ability to infect cultured normal human hepatocytes. ! t appeared that the precore region was dispensable for viral DNA replication, assembly of the mature viral particle and infectivity. Then HBe-minus mutants support, in the absence of helper virus, a complete replication cycle.
iGenotype [~ [ I+lI
] Other combinations
Mild c a l l 4 1 0 4 11
5
Aga~ressiveCAH 3 - - p < .001-
16
3 3 ~ p < .001 --
0
2
In conclusion, it appears that 1) genotypes H is found most frequently in aggressive CAll and the difference with mild CAll is statistically significant (p<.001); 2) in mild CAH genotype I and n are frequently found in asseciation (p<.001 vs aggressive CAH); 3) as duration of disease is not sigaitieanfly different between aggressive and mild CAll but mean age is, it might be hypothesized that the peculiar genutype combination found in mild CAH (I+II) might represent an initial condition before selection of genotype H determined by environmental pressure (40% MURST).
1267
VITAMIN D RECEPTOR GENE POLYMORPHISM AND B O N E LOSS AFTER LIVER TRANSPLANTATION J Guardiola. X X i o l . . I M Eseriba. J M NoUn. E Janrrletta and R Sanie 2
Hospital de Bellvitge y Vilafranca, Barcelona, Spain., Liver Diseases Section 1, NIDDK, NIH, Bethesda, biD, 20892, USA. Background: Bone loss is a frequent complication of orthotopie liver transplantation (OLT). Genetic factors are strong determinants of bone mineral density (BMD). A relationship between common allelie variants in the gene c h i d i n g the vitamin D receptor (VDR) and BMD has recently been demonstrated. Aim: To investigate the influence of VDR gene polymorphism on BMD of cirrhotic patients candidates for OLT and on bone loss after OLT Methods: Fifty six cirrhotic male patients who underwent OLT were studied. Vitamin D receptor polymorphisms were assigned blind with respect to the BMD data following restriction enzyme (Taql, Apal and Bsm) digestion of PCR amplified DNA extracted from formalin fixed tissue. BMD was measured by dual energy x-my absorptiometi'y (DEXA) at the lumbar spine before OLT and at 3 mo (n--37), 6 mo (n=34) and at 12 mo (n=21) after OLT. Values of BMD are reported as Z score (standard deviations from the age and sex adjusted mean). Changes of BMD are expressed as percentage of the initial value. Results: Genotype bb was detected in 22 patients (39%), Bb in 27 (48) and BB in 7 (13%). Before OLT there were no differences between Z scores of bh patients compared to other genotypes (bb: -0.517% vs. Bb/BB: 0.712%, p=n.s.). At 3 mo post OLT the reduction in BMD for bb patients was -1.13% but was -4.57% for Bb and BB cases (13--0.04). By 6 and 12 mo BMD had increased in all genotypes with larger increases seen in bb cases. At 6 mo (bb: +0.37%, Bb/BB: -3.03%, p=0.12) and at 12 mo (bb:+4.55%, Bb/BB -0.45%, p=0.16), receptively. Conclusions: In male patients, VDR gene polymorphisms influence BMD changes after OLT. VDR genotypo bb confers some protection against the rapid bone loss seen in the first six months after OLT.
1268
SELECTIVE I N T E S T I N A L D E C O N T A M I N A T I O N WITH TRIMETHOPRIM-SULFAMETHOXAZOLE IN AN EXPERIMENTAL MODEL OF CIRRHOSIS IN RATS. C Guarner. BA Runvon. S Young. M Heck. and MY Sheikh. Dept. of Medicine. University of Louisville, Louisville, KY. Background: Selective intestinal decontamination with norfloxacin is effective in preventing a) spontaneous bacterial peritonitis (SBP) in cirrhotic patients and b) SBP and bacterial transloeation (BT) due to gram-negative bacteria in a model of cirrhosis in rats (Hepatology 1995; 21:1719). However, norfloxacin increases BT and SBP due to gram-positive bacteria in cirrhotic rats and leads to selection of resistant bacteria which can cause subsequent infection. Therefore, alternative therapies are needed. Trimethoprim-sulfamethoxazole (rMP/SMX) has in vitro activity against gram-negative and some gram-positive bacteria and has been successfully used for preventing bacterial infections in cirrhotic patients (Ann Intern Med 1995,122:595). Aim: To study the efficacy of TMP/SMX in preventing BT and SBP in carbon tetrachloride (CC14) induced cirrhosis in rats. Methods: CCI4 was given to 60 male Sprague-Dawley rats as previously reported (Gastroenterology 1991;100:1737). After 8 weeks of CC14 dosing, half of the rats were treated daily with oral TMP/SMX (20 mg/kg) and half were not. Sacrifice was carried out when the rats were so ill that death was imminent. Mesenteric lymph nodes and ascitic fluid were cultured at the time of sacrifice. Student t and chi-square tests with Yates' correction were used for comparisons and survival curves Were analyzed by Kaplan-Meier method and log-rank test. Results: The development of ascites was delayed (15.3+1.7vs10.4__+0.5 weeks, p<0.05) and there was a trend to longer survival in the treated group. The incidence of BT of the rats that were sacrificed before death was also lower in treated rats than in non-treated (5/10vsll/ll, p<0.05) and there was a trend to less SBP (3/10vs7/ll, pNS). Conclusions: 1) TMP/SMX delayed ascites formation and there was a trend to prolonged survival, 2) TMPISMX reduced the incidence of BT in this experimental model of cirrhosis in rats and there was a trend to less SBP. These results suggest that TMP/SMX may be a good alternative to norfloxacin for preventing SBP.
1265
423A
AASLD ABSTRACTS
HEPATITIS B VIRUS VARIANTS WITH DEFECTS IN THE PRECORE REGION ARE INFECTIOUS IN VITRO. 1phiLinne Gring~, 2,3Shu-Ping Tong. 2Christian Treoo. and 1Christiane Guguen-Guillouzo
1266
1 Unit6 de recherche h6patologique U 49, Institut national de la sant6 et de la recherche m6dicaie, H6pital de PontchaiUou, avenue Henri le Guillou, 35053 Rennes Cedex, 2 Unit6 de recherche sur les h6patites, le SIDA et les r6trovirus humains U 271, Institut national de la sant6 et de la recherche m6dieale, 151, cours Albert Thomas 69424 Lyon Cedex 03, France, 3 Present address: Molecular Hepatology Lab, Massachusets General Hospital Cancer Center, Charlestown, MA 02129.
MILD OR AGGRESSIVE CHRONIC ACTIVE HEPATITIS (CAll) DUE TO HEPATITIS C VIRUS (HCV) ARE ASSOCIATED WITH DIFFERENT COMBINATIONS OF HCV GENOTYPES. ~,~derilflfi.I.Ferretti.A.Ferrari.P .Trande.R.Cosenza.F.Callea*.F .Maaenti.E.Villa. Gastroenterology, Univ.Modena *Spedali Civili,Brescia,Italy. CAH due to HCV infection is o~ten characterized histologically either by very mild inflammatory condition or by a very aggressive pioture, without intermediate situations. As the clinically these patients do not differ significantly, we investigated the details of HCV infection (levels of viremia and genotypes). We studied 52 consecutive patients who had the first recogaitiun of altered ALTs in the previous year, all with histological diagnosis of CAll: 24 had a very mild or borderline CAll (Kundell's score <5) and 27 a n aggressive hepatitis (Knodell's score >9). Viremia was investigated by RT/PCR with endpoint dilutions, HCV genotype by nested PCR according to Okamoto. Biochemical data were not significantly different in the two groups; mean age in mild CAH was significantly younger in comparison with the aggressive CAH (32+10 vs 46_+9). Viremia was not significantly different (mild CAH vs aggressive CAll: 106+102 vs 105+102). HCV genotypes were as follows:
HBV variants defective for e antigen secretion have been frequendy evidenced in blood samples. These variants show several types of mutations, leading to creation of a stop codon or elimination of the initiation codon in the precore region. To determine whether such mutants were still capable of supporting a complete replication cycle we have used an in vitro test system. Mutant viral DNAs bearing a representative panel of mutations in the precore region were transfected in HepG2 cells. The viral particles secreted in the supernatant were analyzed and further tested for ability to infect cultured normal human hepatocytes. ! t appeared that the precore region was dispensable for viral DNA replication, assembly of the mature viral particle and infectivity. Then HBe-minus mutants support, in the absence of helper virus, a complete replication cycle.
iGenotype [~ [ I+lI
] Other combinations
Mild c a l l 4 1 0 4 11
5
Aga~ressiveCAH 3 - - p < .001-
16
3 3 ~ p < .001 --
0
2
In conclusion, it appears that 1) genotypes H is found most frequently in aggressive CAll and the difference with mild CAll is statistically significant (p<.001); 2) in mild CAH genotype I and n are frequently found in asseciation (p<.001 vs aggressive CAH); 3) as duration of disease is not sigaitieanfly different between aggressive and mild CAll but mean age is, it might be hypothesized that the peculiar genutype combination found in mild CAH (I+II) might represent an initial condition before selection of genotype H determined by environmental pressure (40% MURST).
1267
VITAMIN D RECEPTOR GENE POLYMORPHISM AND B O N E LOSS AFTER LIVER TRANSPLANTATION J Guardiola. X X i o l . . I M Eseriba. J M NoUn. E Janrrletta and R Sanie 2
Hospital de Bellvitge y Vilafranca, Barcelona, Spain., Liver Diseases Section 1, NIDDK, NIH, Bethesda, biD, 20892, USA. Background: Bone loss is a frequent complication of orthotopie liver transplantation (OLT). Genetic factors are strong determinants of bone mineral density (BMD). A relationship between common allelie variants in the gene c h i d i n g the vitamin D receptor (VDR) and BMD has recently been demonstrated. Aim: To investigate the influence of VDR gene polymorphism on BMD of cirrhotic patients candidates for OLT and on bone loss after OLT Methods: Fifty six cirrhotic male patients who underwent OLT were studied. Vitamin D receptor polymorphisms were assigned blind with respect to the BMD data following restriction enzyme (Taql, Apal and Bsm) digestion of PCR amplified DNA extracted from formalin fixed tissue. BMD was measured by dual energy x-my absorptiometi'y (DEXA) at the lumbar spine before OLT and at 3 mo (n--37), 6 mo (n=34) and at 12 mo (n=21) after OLT. Values of BMD are reported as Z score (standard deviations from the age and sex adjusted mean). Changes of BMD are expressed as percentage of the initial value. Results: Genotype bb was detected in 22 patients (39%), Bb in 27 (48) and BB in 7 (13%). Before OLT there were no differences between Z scores of bh patients compared to other genotypes (bb: -0.517% vs. Bb/BB: 0.712%, p=n.s.). At 3 mo post OLT the reduction in BMD for bb patients was -1.13% but was -4.57% for Bb and BB cases (13--0.04). By 6 and 12 mo BMD had increased in all genotypes with larger increases seen in bb cases. At 6 mo (bb: +0.37%, Bb/BB: -3.03%, p=0.12) and at 12 mo (bb:+4.55%, Bb/BB -0.45%, p=0.16), receptively. Conclusions: In male patients, VDR gene polymorphisms influence BMD changes after OLT. VDR genotypo bb confers some protection against the rapid bone loss seen in the first six months after OLT.
1268
SELECTIVE I N T E S T I N A L D E C O N T A M I N A T I O N WITH TRIMETHOPRIM-SULFAMETHOXAZOLE IN AN EXPERIMENTAL MODEL OF CIRRHOSIS IN RATS. C Guarner. BA Runvon. S Young. M Heck. and MY Sheikh. Dept. of Medicine. University of Louisville, Louisville, KY. Background: Selective intestinal decontamination with norfloxacin is effective in preventing a) spontaneous bacterial peritonitis (SBP) in cirrhotic patients and b) SBP and bacterial transloeation (BT) due to gram-negative bacteria in a model of cirrhosis in rats (Hepatology 1995; 21:1719). However, norfloxacin increases BT and SBP due to gram-positive bacteria in cirrhotic rats and leads to selection of resistant bacteria which can cause subsequent infection. Therefore, alternative therapies are needed. Trimethoprim-sulfamethoxazole (rMP/SMX) has in vitro activity against gram-negative and some gram-positive bacteria and has been successfully used for preventing bacterial infections in cirrhotic patients (Ann Intern Med 1995,122:595). Aim: To study the efficacy of TMP/SMX in preventing BT and SBP in carbon tetrachloride (CC14) induced cirrhosis in rats. Methods: CCI4 was given to 60 male Sprague-Dawley rats as previously reported (Gastroenterology 1991;100:1737). After 8 weeks of CC14 dosing, half of the rats were treated daily with oral TMP/SMX (20 mg/kg) and half were not. Sacrifice was carried out when the rats were so ill that death was imminent. Mesenteric lymph nodes and ascitic fluid were cultured at the time of sacrifice. Student t and chi-square tests with Yates' correction were used for comparisons and survival curves Were analyzed by Kaplan-Meier method and log-rank test. Results: The development of ascites was delayed (15.3+1.7vs10.4__+0.5 weeks, p<0.05) and there was a trend to longer survival in the treated group. The incidence of BT of the rats that were sacrificed before death was also lower in treated rats than in non-treated (5/10vsll/ll, p<0.05) and there was a trend to less SBP (3/10vs7/ll, pNS). Conclusions: 1) TMP/SMX delayed ascites formation and there was a trend to prolonged survival, 2) TMPISMX reduced the incidence of BT in this experimental model of cirrhosis in rats and there was a trend to less SBP. These results suggest that TMP/SMX may be a good alternative to norfloxacin for preventing SBP.