Clinical associations and prognostic significance of serum anti-p53 antibodies in Thai patients with hepatocellular carcinoma.

Mutations of the p53 gene have been reported to be of prognostic significance in hepatocellular carcinoma (HCC). However, the clinical associations and prognostic value of anti-p53 antibodies, known to be products of the host immune response to these mutations, have been controversial. Serum anti-p53 antibodies were measured in 121 Thai patients diagnosed with HCC using a specific enzyme-linked immunosorbent assay (ELISA) kit. The clinical/pathological characteristics of the patients were compared with respect to the presence of serum anti-p53 antibodies. Cox regression analysis was performed to assess factor interaction and association with survival. Anti-p53 antibodies were detected in 13.2% (16 of 121) of our patients. There were no differences between groups with regard to age, sex, viral markers (HBsAg or anti-HCV), severity of liver disease and tumor advancement. The median survival rates for patients positive and negative for anti-p53 antibodies were 4.0 and 3.0 months, respectively (p = 0.443, by log-rank test). Multivariate analysis demonstrated that an advanced Okuda stage, lack of therapy and presence of portal vein thrombosis were independent factors related to the prognosis of the patients. Nonetheless, the presence of anti-p53 antibodies did not constitute a predictive variable associated with a poorer prognosis. Serum assay of anti-p53 antibodies, although rapid and easily performed, may not be suitable as an alternative to molecular detection of mutations in assessing tumor advancement and prognosis of patients with HCC.

Increasing susceptibility to HAV among members of the young generation in Thailand.

The prevalence of antibodies to hepatitis A virus was studied in 961 children and adolescents, randomly selected from five different provinces in Thailand (Chonburi, Lopburi, Udonthani, Nakhon Si Thammarat and Lopburi). The highest prevalence was found in Nakhon Si Thammarat, with 32.1 percent of those aged 10-14 years and 57.1 percent of those aged 15-18 years showing evidence of protective immunity. However, this high rate could be explained by an outbreak of hepatitis A in 1992. In the remaining four provinces, the pattern was typically age-related in that all individuals showed between zero and 13 percent antibody prevalence until reaching the 15-to-18-year age group where it increased to between 5.6 and 22.7 percent. The overall sero-prevalence among all age groups was 7.9 percent. Thus, the majority of the younger generation is susceptible to hepatitis A virus infection thereby enhancing the impact, should an outbreak occur. Preventive measures that might be taken are education aimed at better hygiene and sanitation, as well as vaccination of susceptible individuals within high-risk populations.

Core promoter and precore mutants of hepatitis B virus: prevalence and clinical relevance in chronic hepatitis patients.

The present study was conducted to determine prevalence and exact type, as well as nucleotide position of the precore/core mutations of hepatitis B virus found in Thai patients diagnosed with chronic hepatitis and/or cirrhosis in relation to the clinical parameters established with the respective patients. To that end, 24 HBeAg-positive and 56 HBeAg-negative individuals were selected at random from a cohort of altogether 256 chronic liver disease patients. DNA was extracted from their blood sera, amplified by polymerase chain reaction using semi-nested primers and subjected to direct sequencing. Clinically, the HBeAg-positive chronic hepatitis patients displayed significantly higher transaminase levels than those negative for HBeAg. Our results showed 2 of the 7 (28.6%) PCR-positive HBeAg-positive sera displaying double mutations in the core promoter region at position 1762/64. The nucleotide sequences obtained from the 24 PCR-positive HBeAg-negative sera revealed 18 (75%) mutations in the core promoter region (1762/64), and/or 7 (29.2%) mutations at position 1753, and/or 6 (25%) mutations of the start codon (1814), and/or 8 of (33.3%) nucleotide 1896 turning codon 28 into a stop codon and one sample (4.2%) displaying a deletion between nucleotides 1758-1772. It is suggested that the mutations observed have an impact on the DNA secondary structure in such a way that successful transcription of the HBeAg gene is rendered impossible. To what extent this mutation influences the severity of chronic liver disease remains to be elucidated.

Parvovirus B19 antibodies in immunocompromized children in Thailand.

Parvovirus B19, a non-enveloped single stranded DNA virus is distributed worldwide. Sero-prevalence in adult populations amounts to approximately 50%. Clinical manifestations vary depending on the Immune status of the infected individuals and may include mild childhood Infection as well as hydrops fetalis due to intrauterine infection. To determine the prevalence of this infection among the immunocompromized individuals in Thailand, we determined, by indirect ELISA, levels of IgM and IgG antibodies to the parvovirus B19 in 106 immunocompromized children. These included 49 children who were on chemotherapy for treatment of malignancies, 18 who were receiving immunosuppressive drugs after organ transplantations, 14 who were under a regimen of corticosteroids and 25 who were positive for antibodies to HIV. The average prevalence of IgG antibodies in 106 children was 16.0%; the prevalence of antibodies was 33.3% in post-transplanted group, 16% in children positive for HIV, 12.2% in the group receiving chemotherapy for malignancies and 7.6% in the group treated with corticosteroids. All children were negative for IgM antibodies to parvovirus B19.

Prevalence of antibodies to parvovirus B 19 in Thailand.

Infection with human parvovirus B 19, a single-stranded non-enveloped DNA virus of worldwide distribution, is rather common and displays a broad spectrum of clinical manifestations of varying severity, depending on the patient's immune response. As the target of infection are the erythroid precursor cells, patients can experience an aplastic crisis. Usually, at least in immunocompetent individuals, viremia ceases with the appearance of virus-specific antibodies in the patient's serum whereupon the patients retain lifelong immunity to reinfection. Since data as to the prevalence of this agent has not been established for Thailand, the purpose of the present study was to investigate its frequency among 3 distinct groups, comprising 30 healthy children. 64 children with acute unrelated illness, and 35 voluntary blood donors, respectively, by means of enzyme linked immunosorbent assay. Our results have shown that, as reported for other countries, anti-parvovirus IgG increases in an age-dependent manner and is established at an overall prevalence of 20.16%, inviting the conclusion that the local population is infected by this agent as frequently as those of other countries in the Far East. Further studies need to be undertaken in order to elucidate its prevalence among members of high-risk groups.

Cytomegalovirus infection in immunocompromised children in Thailand.

Cytomegalovirus (CMV) constitutes the most widespread cause of congenital and perinatal viral infections on a global scale, exceeding a 90%-prevalence among blood donors in Thailand. The present study was aimed at determining the prevalence of CMV in the sera of 113 immunocompromised children by means of serology, as well as polymerase chain reaction using nested primers. Our results showed anti-CMV IgG, i.e. latent infection, prevalent in an age-dependent manner irrespective of the disorder underlying the children's immunocompromised condition, whereas anti-CMV IgM was equally prevalent throughout all age groups and disease patterns and, therefore, unreliable as a marker. Detection of serum CMV DNA by PCR represented the most exact diagnostic parameter by far, indicating active infection long before clinical symptoms may appear. In conclusion, based on the high prevalence of latent CMV infection among the general population in Thailand, we recommend especially the sera of immunocompromised patients be examined for the presence of viral DNA by means of PCR in order to provide clinical guidelines for their proper management.

Coinfections with hepatitis g and/or c virus in hepatitis B-related chronic liver disease.

Concurrent infections with HGV and/or HCV (HGV/HCV) were investigated in 196 patients with HBV-related chronic liver disease (115 chronic hepatitis, 31 liver cirrhosis, 50 hepatocellular carcinoma), and in 100 HBsAg carriers. Coinfections were detected in 18 (9.2%) patients with HGV (10) or HCV (5) or both agents (3), but in none of the HBsAg carriers. Patients with coinfection were more frequently exposed to blood transfusions (55.6% vs 5.6%) and also were more commonly anti-HBe positive. Serum levels of HBV-DNA were lower in patients with HCV coinfection than in those coinfected with HGV. Interferon was administered to 39 patients with chronic active hepatitis including 7 patients with HGV/HCV coinfection. Sustained clearance of HBV-DNA was observed in 10 (25.6%) patients who were solely infected with HBV. These patients were significantly younger and had much lower histological scores than non-responders. Patients with HCV coinfection had significantly higher pre-treatment histological scores than those without HCV. After interferon treatment, a significant reduction in histological scores was observed in all patients except those coinfected with HGV/HCV. None of the 7 patients with coinfection had sustained clearance of HBV-DNA or HCV-RNA, and only one had cleared HGV-RNA. These results suggest that parenteral exposure is a risk factor for HGV/HCV coinfection in chronic HBV infection. HGV infection shows no significant impact on chronic HBV infection. HCV coinfection appears to inhibit HBV replication, but causes more severe chronic hepatitis and increases resistance to interferon therapy.

Prevalence of core promotor and precore mutants of hepatitis B virus in thailand by RFLP and sequencing.

Hepatitis B virus has been known to frequently undergo mutations of its genome at various sites, mostly due to it employing a reverse transcriptase devoid of proofreading capacity in the course of its replication. The purpose of the present study has been to screen 257 HBsAg-positive chronic liver disease patients, more specifically 78 cases chosen at random out of those negative for HBeAg and 33 of the HBeAg-positive cases serving as controls for three discreet point mutations in the precore/core region of hepatitis B virus. To that end, HBV DNA extracted from sera was amplified by polymerase chain reaction (PCR) using semi-nested primers and subsequently subjected to restriction fragment length polymorphism (RFLP) analysis, 36 HBeAg-negative versus 30 HBeAg-positive sera, respectively, as well as to direct sequencing in some samples randomly selected to corroborate the RFLP results. Our results showed double mutations at positions 1762 and 1764 of the core promoter in between 25/36 (69.4%) and 19/25 (76%) of the sera tested, a missense mutation of the start codon in between 8/36 (22.2%), and 5/25 (20%) and a mutation turning codon 1896 into a stop codon in between 9/36 (25%) and 6/25 (24%) determined by RFLP and sequencing, respectively. These data indicate the double mutation at positions 1762 and 1764 to be the most prevalent among HBeAg-negative chronic hepatitis patients in Thailand whereas, in contrast to reports from other Asian countries, the mutation at nucleotide 1896 occurred in a mere 25%, while on the other hand the mutation abolishing the start of protein synthesis was observed to occur at a higher frequency than determined in several other geographical areas.

Hepatitis viruses and chronic liver disease.

We have investigated several groups of Thai patients diagnosed with chronic liver disease including chronic hepatitis, cirrhosis and hepatocellular carcinoma, as well as cholangiocarcinoma, for the prevalence of infection with either one of the hepatitis viruses B, C, G and the novel hepatitis virus TT (TTV). The 168 patients tested comprised 120 men and 48 women with their median age ranging from 42.3 to 62.3 years. Screening for antibodies to HBV and HCV was performed by a commercially available serological test kit, for the presence of HBV and TTV DNA by PCR, and of HCV and HGV RNA by RT-PCR, respectively. There was a clear two-fold higher prevalence of HBV (49%) over HCV (27%) infection and a four-fold higher frequency compared to HGV (13%) and TTV (11%) infection, respectively, in those individuals with chronic hepatitis, cirrhosis, and hepatocellular carcinoma, whereas all but one patient with cholangiocarcinoma the etiology of which has been ascribed to parasitic infestation, were free of all viral markers. In Thailand chronic HBV, and to a lesser extent, chronic HCV infection represent the two most common causes of hepatitis potentially proceeding to chronic liver disease, whereas the clinical significance pertinent to HGV and TTV remains to be elucidated.

TT virus infection in intravenous drug users.

Our group has investigated 201 intravenous drug users for the presence of TTV DNA by means of polymerase chain reaction (PCR). The majority of the individuals tested were male, their age ranging from 16 to 63 years, and the duration of intravenous drug use from one to 40 years. TTV DNA was present in 62 of the 201 IVDUs (30.8%) with its prevalence on the ascent between the age groups below 20 and those between 21 and 30 years, as well as between the groups below 60 and between 60 to 120 months' duration of drug intake, respectively. When tested again after 9 months, nine IVDU (23.7%) were found TTV negative by PCR hinting at potential immunological clearance. Our control group comprised 200 healthy blood donors, 7% of whom were found to harbor TTV DNA in an age-dependent fashion, as observed with the IVDU. From the liver function tests performed we could not detect any statistically significant difference regarding ALT elevation observed in TTV-positive compared with TTV-negative individuals. To date, TTV does not appear to cause any serious liver disease in the majority of cases examined.

Clearance of hepatitis TT virus infection among thalassemia children and IVDU.

The novel transfusion transmissible hepatitis virus TTV first isolated by a group from Japan has predominantly been detected in members of groups at high risk for contracting blood borne viruses. Aside from elevated liver enzymes, the symptoms associated with its infection have been reported to range from asymptomatic to hepatic failure. The purpose of the present study was to determine if and to what extent the host's immune response is capable of clearing TTV infection. Hence, we extracted DNA from sera obtained from altogether 201 intravenous drug users (IVDU) and 80 thalassemia children--both groups at high risk of parenteral exposure--and performed PCR using semi-nested primers. Those positive for TTV DNA were once again subjected to PCR after approximately one year in order to determine how many still harbored the virus. Our results showed TTV DNA to be absent in merely 20.6% of the formerly positive IVDU, whereas it was still present in all the thalassemia children who could be tested for the second time. Based on the small sample size and the high-risk environment, these results ought to be interpreted with caution and definitely merit further investigation.

Susceptibility to hepatitis A virus infection among chronic liver disease patients and healthy blood donors in Thailand.

Due to improvements in socio-economic and sanitation conditions, Thailand has undergone a change from hyperendemicity to intermediate endemicity for hepatitis A virus infection, leaving a large part of the adult population without immunity. At the same time, the country is still highly endemic for hepatitis B and especially in the northeast, hepatitis C virus infection both of which when acquired during infancy or early childhood exhibit a strong tendency to turn towards chronic liver disease, although in particular with hepatitis B virus the asymptomatic carrier state is also rather common. As no cross-immunity exists between any of these viruses, double or triple infections do occur, a situation where previously acquired immunity to HAV becomes crucial as double infections have been shown to take a more severe or even fatal course. In the present study, we investigated 820 HBV- and/or HCV-related chronic liver disease (CLD) patients and 195 blood donors, both groups divided by 10-year age intervals, for the prevalence of anti-HAV. The results showed the same age dependence of immunity for all groups tested as can be expected for an area of intermediate endemicity, in that approximately 50% of those between 21 and 30 years of age had acquired anti-HAV. These findings indicate the immune response to HAV infection not to be altered by chronic infection with either HBV or HCV. Hence, vaccination against HAV should be considered, particularly in anti-HAV-negative patients with CLD.

Does hepatitis G virus cause significant clinical liver disease?

Regarding the newly discovered hepatitis G virus (HGV), little is known about its relation to the cause and clinical significance of acute and chronic liver disease and hepatocellular carcinoma. Lacking a reliable serum immunoassay, the only method available for detecting the viral RNA in patients consists of the rather costly and time consuming RT-PCR. HGV has a worldwide distribution with up to 5% voluntary and 12.9% commercial blood donors infected, yet it appears to be asymptomatic. Moreover, HGV is frequently found as a coinfection with HCV or, to a lesser extent, HBV with symptoms tending to follow the patterns known for HCV or HBV infection, respectively. Being a blood-borne virus, it is most prevalent among members of high risk groups, such as IVDUs, patients on hemodialysis, recipients of blood and blood products and patients infected with HCV, HBV, or HIV, HGV can be parenterally, vertically, or sexually transmitted and after prolonged exposure, the virus may be eliminated by the patient's immune response. As yet, no unambiguous evidence exists regarding HGV's role in causing acute or chronic liver disease and, apart from a few isolated reports to the contrary, the infections appear rather mild. Therefore, more studies are required before a decision can be made whether to routinely screen blood donors for the presence of HGV RNA.