Detection of hepatitis C virus RNA by in situ hybridization in paraformaldehyde fixed biopsies.

Fourteen hepatitis C virus (HCV) chronically infected patients were submitted to routine liver biopsy for histological evaluation. Liver samples were assayed to HCV-RNA by in situ hybridization, using digoxigenin labeled probe. HCV genotypes were found to be predominantly type 1 (71.4%), followed by genotype 3 (21.4%), and genotype 2 (7.2%). Alanine-aminotransferase levels were raised in 10 patients. The histopathological scores were minimal (21.4%), mild (57.2%), and moderate (21.4%). Viral RNA was detected in liver cells from nine patients (64.3%). ISH method provides localization and poor confirmation of HCV RNA in the liver tissue of HCV chronic patients.

Detection of hepatitis C virus RNA by in situ hybridization in paraformaldehyde fixed biopsies

Fourteen hepatitis C virus (HCV) chronically infected patients were submitted to routine liver biopsy for histological evaluation. Liver samples were assayed to HCV-RNA by in situ hybridization, using digoxigenin labeled probe. HCV genotypes were found to be predominantly type 1 (71.4 percent), followed by genotype 3 (21.4 percent), and genotype 2 (7.2 percent). Alanine-aminotransferase levels were raised in 10 patients. The histopathological scores were minimal (21.4 percent), mild (57.2 percent), and moderate (21.4 percent). Viral RNA was detected in liver cells from nine patients (64.3 percent). ISH method provides localization and poor confirmation of HCV RNA in the liver tissue of HCV chronic patients.

Experimental hepatitis A virus (HAV) infection in Callithrix jacchus: early detection of HAV antigen and viral fate.

Common marmosets (Callithrixjacchus) were orally inoculated with a Brazilian strain (HAF-203) of hepatitis A virus (HAy). Three monkeys were euthanized at postinoculation hours 6, 12 and 24 to investigate the early events of HAV infection. Following others three inoculated and one control marmosets remained throughout the 46 day to evaluation of viral excretion. Different samples were collected to detect sequential presence of HAV RNA by nested reverse transcription-polymerase chain reaction (RT-PCR) in liver, saliva, bile and stools at 6 hours to 461h days postinoculation. Liver tissues were examined by immunofluorescence assay in a confocal laser-scanning microscope for the presence of HAV antigen. HAV RNA was detected in saliva during the course of the study, in bile from 24 hours to 46 days. in stools from 7 to 46 days and liver at 12 hours postinfection. In immunofluorescence of liver stained preparations, viral antigen was present at six hours after inoculation throughout the remainder of the 46-day study. The animals developed histological and biochemical acute hepatitis after second week postinoculation. Spleen, duodenum, and mesenteric lymph nodes specimens were negative for HAV antigens. This study supports the possibility that in Callithrixjacchus orally inoculated with hepatitis A virus the saliva route may be additional way of viral elimination. The viral replication in the liver was responsible for biliary HAV presence and latter HAV detection in fecal samples.

Seprafilm reduces adhesions to polypropylene mesh.

BACKGROUND: Adhesions to polypropylene mesh used for abdominal wall hernia repair may eventuate in intestinal obstruction or enterocutaneous fistula. A Seprafilm Bioresorbable Membrane translucent adhesion barrier has been shown to inhibit adhesions. This investigation was designed to determine if Seprafilm alters abdominal visceral adhesions to polypropylene mesh. METHODS: A 2.5-cm square abdominal muscle peritoneal defect was created and corrected with polypropylene mesh. Mesh alone was used in 17 rats. In another 17, the Seprafilm membrane was applied between the viscera and the mesh. Five animals had the bioresorbable membrane placed in the subcutaneous space and between the mesh and the viscera. Laparoscopy was performed 7, 14, and 28 days later to evaluate adhesions as a percentage of mesh surface involved. RESULTS: Polypropylene mesh alone was associated with adhesions in every rat. The average area involved was 90%, the minimum was 75%. Adhesions were present within 24 hours and progressed up to 7 days with no change thereafter. When the Seprafilm barrier was used, the mean area involved was 50%. In 16 such rats, the area involved was smaller than any control animal. No adhesions formed in 5 animals. Scanning electron microscopy demonstrated a mesothelial cell layer covering the mesh after 4 weeks. CONCLUSIONS: The use of the Seprafilm adhesion barrier resulted in a significant reduction of adhesion formation to polypropylene mesh (P <.001).

Inducible nitric oxide synthase (iNOS) expression in liver and splenic T lymphocyte rise are associated with liver histological damage during experimental hepatitis A virus (HAV) infection in Callithrix jacchus.

Callithrix jacchus is considered a reliable animal model for hepatitis A virus (HAV) infection. All three HAV orally inoculated marmosets developed hepatitis - the infection was monitored by continuous virus shedding, high levels of serum enzyme alanine aminotransferase, specific antibody and seroconversion 3-6 weeks after HAV inoculation. HAV antigen was detected in liver by immunofluorescence 4 days post inoculation (PI) and onwards. To gain insight into the biological role of inducible nitric oxide synthase (iNOS) during immune-related acute liver injury the enzyme was searched in frozen biopsies: immunofluorescent labeling was found in the cytoplasm of liver cells mainly Kupffer's cells and spleen macrophages (CD68+) starting 11 days PI with maximum intensity on the fifth to sixth week PI. Necroinflammatory liver lesions characteristic of viral hepatitis were also observed at 10 days PI with maximum severity at 4 to 6 weeks PI. Furthermore, T lymphocytes (CD2+) were raised at this time point. No difference was evident in the frequency of B lymphocytes (CD20+). Therefore, iNOS expression preceded necroinflammatory liver lesion and maximal immunofluorescence reaction was coincident with tissue injury, supporting the hypothesis that NO contributes to hepatic cytotoxic mechanism but also to virus clearance. The concomitant rise in T-lymphocyte population may suggest a role for these cells in this and/or other independent HAV-induced pathological changes.

Abdominal adhesions to prosthetic mesh evaluated by laparoscopy and electron microscopy.

BACKGROUND: Most adhesion experiments involve observations at a single time point. We developed a method to evaluate abdominal adhesions to surgical mesh by sequential laparoscopy. STUDY DESIGN: An abdominal wall defect was created in rats and repaired with polypropylene mesh. Sequential laparoscopic evaluation of adhesion formation was performed in each animal. The percentage of mesh area involved was scored (0% to 100%). At various time intervals animals were sacrificed and samples were obtained for light and scanning electron microscopy. RESULTS: Adhesions were already present on day 1, increased by day 7, and did not progress thereafter. Mesh surfaces free of adhesions were covered with a confluent mesothelial cell layer, first seen by scanning electron microscopy on day 5 and complete by day 7. CONCLUSIONS: Intraabdominal adhesions are best studied by sequential laparoscopy. Adhesions develop within 1 day of prosthesis placement. Adhesion-free surfaces are carpeted with mesothelial cells by day 7 and remain free thereafter, for duration of study.

Radioprotection of the intestinal mucosa of rats by probucol.

Probucol is a lipid-regulating drug that also has antioxidant properties. This study was designed to test the possibility that probucol could provide radioprotection of the intestine when administered either intralumenally or systemically. Tissue damage was evaluated histologically by quantifying the number of crypts per circumference and the mucosal height. Animals were sacrificed 5 days after 11 Gy of X irradiation. In one series of experiments, a loop of mid small bowel was exteriorized operatively and compartmentalized into segments, each filled with probucol or saline. Intralumenal administration of probucol prior to irradiation led to a significantly greater number of crypts per circumference and mucosal height compared to saline-filled irradiated controls. In another series of experiments, five groups of rats were irradiated: (1) probucol in the small bowel lumen, (2) intravenous probucol, (3) probucol by gavage, (4) probucol added to standard rat chow and (5) saline control. In the rats given probucol intravenously prior to X irradiation, crypt numbers and mucosal height were significantly enhanced. Probucol given by gavage also resulted in protection. Rats fed a diet containing probucol showed no significant protection. Topical administration was more effective than systemic. Probucol protects the intestinal mucosa from acute radiation damage when given topically, intravenously or by gavage, but does not do so when given as a dietary supplement.

Hepatitis B epidemiology and cultural practices in Amerindian populations of Amazonia: the Tupí-Mondé and the Xavánte from Brazil.

Hepatitis B infection and disease are highly endemic in South America. Prevalences of positivity are particularly high in Amazonia, and among Amerindian peoples in particular. This paper reports the results of a seroepidemiological survey for hepatitis B virus (HBV) carried out among four Amerindian populations from the Brazilian Amazon region: Gavião, Surui, Zoro and Navate. Rates of positivity to HBV serological markers (HBsAg, anti-HBs and or anti-HBc) are very high for the four groups, ranging from 62.8 to 95.7%. It is argued that the high rates of positivity in the Amerindian groups dealt with in this study, as well as for other Amazonian populations, are related to a complex of cultural practices which enhance the likelihood of HBV transmission (bloodletting, scarification, tattooing and orally processed food, among others). The authors suggest that, due to unique patterns of interaction between sociocultural and environmental factors. HBV infection assumes a specific profile in native Amazonian societies.

Intestinal radioprotection by vitamin E (alpha-tocopherol).

OBJECTIVES: The major objective of this study was to test vitamin E as a potential radioprotectant for the small bowel of the rat. SUMMARY BACKGROUND DATA: Vitamin E has previously been shown to provide radioprotection in animal models: increased survival after whole-body irradiation, diminished absorptive malfunction, and modest diminution in postirradiation hemolysis. The lumenal route for intestinal radioprotection has not been tested. METHODS: Rat mid-small bowel was surgically exteriorized and segmented by ties into compartments, each of which was filled with a test solution 30 minutes before 1100 cGy of x-irradiation was administered. After the rats were killed 5 days later, the various segments were evaluated for surviving crypts, mucosal height, and goblet cell preservation. Lumenal agents included alpha-tocopherol phosphate and alpha-tocopherol acetate. In a separate study, dietary supplements of alpha-tocopherol were given for 10 days before irradiation, and the same irradiation sequence was carried out. RESULTS: Small bowel crypt cell numbers, mucosal height, and goblet cell numbers were significantly protected from radiation effects by dietary alpha tocopherol pretreatment and by lumenal application of the vitamin. CONCLUSIONS: These studies indicate that vitamin E can serve as a partial protectant against acute irradiation enteritis, whether given as chronic oral systemic pretreatment or as a brief topical application.

Histopathological and immunohistochemical studies of hepatitis A virus infection in marmoset Callithrix jacchus.

Samples of serum, feces and liver tissue and organs of six cotton-eared marmosets Callithrix jacchus infected intravenously with two different strains of hepatitis A virus (HAV), were studied by conventional histologic techniques, by serological techniques and by immunocytochemical methods, such as immunofluorescence (IF) and peroxidase-antibody techniques. Hepatitis A antigen (HAAg) was detectable in daily collected stools, in liver biopsy obtained sequentially, and in organs collected at necropsy. Two marmosets also developed antibodies to HAV. By contrast, serum transaminases were not altered and there were histological hepatic lesions consistent with acute viral hepatitis in all inoculated animals. The data obtained, demonstrate that these primates are susceptible to human HAV and may be a useful animal model for the study of infection by this virus.

Immunization against hepatitis B in children from endemic zone: evaluation of the antibody response against the DNA recombinant vaccine (Engerix B-20 MCG).

A previous seroepidemiological study in the rural zone of Vargem Alta (ES) SouthEast of Brazil, showed a prevalence of up to 9% of hepatitis B surface antigen (HBsAg) in some areas. One hundred susceptible children aging 1 to 5 years old were selected and immunized with a recombinant DNA hepatitis B vaccine (Smith-Kline 20 mcg) using the 0-1-6 months vaccination schedule. Blood samples were collected at the time of the first vaccine dose (month 0) in order to confirm susceptible individuals and 1,3,6 and 8 months after the first dose, to evaluate the antibody response. Our results showed that two and five months after the second dose, 79% and 88% of children seroconverted respectively, reaching 97% after the third dose. The levels of anti-HBs were calculated in milli International Units/ml (mIU/ml) and demonstrated the markedly increase of protective levels of antibodies after the third dose. These data showed a good immunogenicity of the DNA recombinant hepatitis B vaccine when administered in children of endemic areas.

Detection of carriers of X-linked gene for Duchenne muscular dystrophy by levels of creatine kinase and pyruvate kinase.

Creatine kinase (CK) is the enzyme most often utilized for the detection of carriers of the gene for X-linked muscular dystrophies. In 1974, pyruvate kinase (PK) levels were also found to be increased in these carriers. The objective of the present study, carried out on 77 women related to patients with Duchenne muscular dystrophy (DMD), was to compare the efficiency of the two enzymes in the detection process. Of the 11 obligate heterozygotes for the DMD gene in the group, 8 exhibited elevated mean CK levels, 6 had elevated mean PK levels, and 9 had elevated mean levels of at least one of the enzymes. Among the mothers of isolated patients, 2/13 had elevated mean CK levels, 3/13 had elevated mean PK levels, and 5/13 had elevated mean levels of at least one of these enzymes. Thus, the study confirms data obtained by other investigators indicating that the use of PK can increase the detection rate of carriers of the gene for X-linked muscular dystrophies.