Mutant p53-R273H mediates cancer cell survival and anoikis resistance through AKT-dependent suppression of BCL2-modifying factor (BMF).

p53 is the most frequently mutated tumor-suppressor gene in human cancers. Unlike other tumor-suppressor genes, p53 mutations mainly occur as missense mutations within the DNA-binding domain, leading to the expression of full-length mutant p53 protein. Mutant p53 proteins not only lose their tumor-suppressor function, but may also gain new oncogenic functions and promote tumorigenesis. Here, we showed that silencing of endogenous p53-R273H contact mutant, but not p53-R175H conformational mutant, reduced AKT phosphorylation, induced BCL2-modifying factor (BMF) expression, sensitized BIM dissociation from BCL-XL and induced mitochondria-dependent apoptosis in cancer cells. Importantly, cancer cells harboring endogenous p53-R273H mutant were also found to be inherently resistant to anoikis and lack BMF induction following culture in suspension. Underlying these activities is the ability of p53-R273H mutant to suppress BMF expression that is dependent on constitutively active PI3K/AKT signaling. Collectively, these findings suggest that p53-R273H can specifically drive AKT signaling and suppress BMF expression, resulting in enhanced cell survivability and anoikis resistance. These findings open the possibility that blocking of PI3K/AKT will have therapeutic benefit in mutant p53-R273H expressing cancers.

Field evaluation of a rapid diagnostic test to detect antibodies in human toxocariasis.

Human toxocariasis which is caused mainly by the larvae of Toxocara canis and Toxocara cati, is a worldwide zoonotic disease that can be a potentially serious human infection. The enzyme-linked immunosorbent assay (ELISA) using T. canis excretory-secretory (TES) antigens harvested from T. canis larvae is currently the serological test for confirming toxocariasis. An alternative to producing large amounts of Toxocara TES and improved diagnosis for toxocariasis is through the development of highly specific recombinant antigens such as the T. canis second stage larva excretory-secretory 30 kDa protein (recTES-30). The aim of this study was to evaluate the sensitivity and specificity of a rapid diagnostic kit (RDT, named as iToxocara kit) in comparison to recTES-30 ELISA in Serendah Orang Asli village in Selangor, Malaysia. A total of 133 subjects were included in the study. The overall prevalence rates by ELISA and RDT were 29.3% and 33.1%, respectively, with more positive cases detected in males than females. However, no association was found between toxocariasis and gender or age. The percentage sensitivity, specificity, positive predictive value and negative predictive value of RDT were 85.7%, 90.1%, 80% and 93.2%, respectively. The prevalence for toxocariasis in this population using both ELISA and RDT was 27.1% (36/133) and the K-concordance test suggested good agreement of the two tests with a Cohen's kappa of 0.722, P<0.01. In addition, the followed-up Spearman rank correlation showed a moderately high correlation at R=0.704 and P<0.01. In conclusion, the RDT kit was faster and easier to use than an ELISA and is useful for the laboratory diagnosis of hospitalized cases of toxocariasis.

Lack of islet neogenesis plays a key role in beta-cell depletion in mice infected with a diabetogenic variant of coxsackievirus B4.

Group B coxsackieviruses (CVBs) have a well-established association with type 1 diabetes but the mechanism of depletion of beta-cell mass following infection has not yet been defined. In this report we show that the major difference in pathogenesis between the E2 diabetogenic strain of CVB4 and the prototypic JVB strain in SJL mice is not in tropism for islet cells but in the degree of damage inflicted on the exocrine pancreas and the resulting capacity for regeneration of both acinar and islet tissue by the host. Both strains replicated to a high titre in acinar tissue up to day 3 post-infection (p.i.), while the islets of Langerhans were largely spared. However, the pancreas in the JVB-infected animals then regenerated and many small islets were seen throughout the tissue by day 10 p.i. In contrast, the acinar tissue in E2-infected mice became increasingly necrotic until all that remained by day 21 p.i. were large islets containing varying numbers of dead cells, caught up in strands of connective tissue. Surviving beta cells were found to synthesize little insulin, although islet amyloid polypeptide was detected and glucagon synthesis in alpha cells appeared normal or enhanced. Our results suggest that the key to CVB-E2-induced damage lies in the exocrine tissue and prevention of islet neogenesis rather than from direct effects on existing islets.

Highly unsaturated fatty acid induced tumour regression in glioma pharmacodynamics and bioavailability of gamma linolenic acid in an implantation glioma model: effects on tumour biomass, apoptosis and neuronal tissue histology.

Highly unsaturated fatty acids (HUFAs) are naturally occurring anti-tumour agents. HUFAs act as intracellular signalling molecules in cell proliferation and death. In human glioma, HUFAs may stimulate tumour regression and apoptosis. An implantation glioma model, using the C6 glioma cell line, was used to investigate the bioactivity of locally infused n-6 HUFA gamma linolenic acid (GLA). Rat brains (15 normal and 37 C6 tumour bearing) were infused with vehicle or GLA 200 microM-2 mM. The most active local concentration of GLA for anti-tumour activity was 2 mM, infused at 1 microl/h over 7 days. Tumour regression, increased apoptosis and decreased proliferation were observed in tumours of rats infused with this concentration of GLA. Little effect on normal neuronal tissue was detected. The intraparenchymal route was an effective method of GLA administration in the treatment of glioma. These studies provide further insights into the potential role of HUFAs as anti-glioma agents.

Tropical liver abscess.

Forty-one consecutive cases of liver abscesses seen at the National University Hospital, Singapore from 1988 to 1994 were reviewed. Twenty-seven cases (65%) were pyogenic, six (15%) amoebic, two (5%) tuberculous and six (15%) indeterminate. The predominance of pyogenic abscesses is in marked contrast to previous studies from the region a decade ago in which amoebic abscesses were the commonest type. The commonest pathogen causing pyogenic abscess was Klebsiella pneumoniae. Two cases were due to Mycobacterium tuberculosis, and this organism needs to be actively looked for in smears and cultures of aspirated material. As the majority of organisms isolated were resistant to ampicillin, empirical antibiotic treatment for suspected pyogenic abscess should include gentamicin or a cephalosporin. Percutaneous needle aspiration of the abscess was performed for 85% of pyogenic abscesses and surgery was necessary in only two cases because of complications. We found that percutaneous aspiration of liver abscess is helpful to confirm the diagnosis, provides a better bacteriological culture yield, gives a good outcome, and may uncover clinically unsuspected conditions like malignancy and tuberculoma which may mimic the presentation of liver abscesses. We recommend routine cytological examination of aspirated abscess material as well as stains and cultures for acid-fast bacilli.

Wilson's disease presenting as haemolytic anaemia and its successful treatment with penicillamine and zinc.

Haemolysis is an uncommon first manifestation of Wilson's disease. We describe a young woman who presented with episodic haemolysis and abnormal liver functions; the diagnosis of Wilson's disease was not made until nine months later. She responded well to a combination of penicillamine and zinc. This report underscores the importance of considering Wilson's disease as a cause in a patient with haemolysis of uncertain aetiology, since the disease can be successfully treated in the early stages. the mechanism of oxidative damage to erythrocytes by the excessive copper and the present role of zinc therapy are also discussed.

Subcutaneously administered recombinant human beta-interferon in the treatment of chronic hepatitis B virus infection.

BACKGROUND: Treatment of chronic replicative hepatitis B virus (HBV) infection is aimed at stopping viral replication and preventing the development of chronic liver disease. beta-Interferon treatment has been less well studied than alpha-interferon. METHODS: The efficacy and tolerability of a 6-month course of subcutaneously administered human recombinant beta-interferon (rINF-beta ser) was studied and the results of a low-dose regime compared with a high-dose regime. Twenty patients (17 men and three women), aged 24-54 years, with chronic hepatitis B virus infection (all hepatitis B surface antigen-positive with detectable HBV-DNA in their sera for at least 3 months prior to therapy) were randomized into two treatment groups of 10 patients each. The low-dose group received 6 x 10(6) U/dose and the high-dose group received 30 x 10(6) U/dose, both groups receiving their respective doses three times a week initially for 1 month and continuing for a total of 6 months. RESULTS: The treatment was well tolerated in both groups. None of the patients required dosage reduction or cessation of treatment because of side-effects. HBV-DNA decreased in all patients during treatment, demonstrating the anti-viral efficacy of rINF-beta ser, and was undetectable in 20 and 40% of patients receiving low-dose and high-dose regimes, respectively, at the end of 6 months treatment (P = N.S.). One year after completion of treatment, HBV-DNA was undetectable in 50 and 30% of patients in the low-dose and high-dose groups, respectively (P = N.S.). However, only one patient achieved seroconversion with loss of the hepatitis B surface antigen and appearance of an antihepatitis B 'e' antigen at the end of 18 months. CONCLUSION: This study shows that subcutaneously administered rINF-beta ser is well tolerated, but the optimal dose and duration of treatment still needs to be defined by further studies.

Fine needle aspiration biopsy of small/intermediate cell tumors in the liver. Considerations in a southeast Asian population.

OBJECTIVE: To assess the diagnostic problems and accuracy involved in rendering an exact cytologic diagnosis, including reference to a primary site of origin, on fine needle aspiration biopsies (FNABs) of small/intermediate cell tumors of the liver. STUDY DESIGN: Thirty-five hepatic FNABs of small/ intermediate tumors, neuroendocrine tumors (NETs) and poorly differentiated or undifferentiated cancers occurring in adults were analyzed. Ancillary studies, including immunohistochemistry, were performed whenever necessary. All other relevant histopathologic sections and medical records were reviewed. RESULTS: There were 26 metastases, while 9 were considered primary lesions. The aspirates were categorized into 11 NETs (pancreas 5, lung 1, liver 2, unknown primary 3); 9 undifferentiated nasopharyngeal carcinomas; 2 small cell undifferentiated carcinomas (lung 2); 4 undifferentiated carcinomas, not otherwise specified (lung 1, pancreas 1, liver 1, unknown 1); 1 lobular-ductal carcinoma (breast); 1 glioblastoma multiforme (brain); 1 ovarian carcinoma; 1 non-Hodgkin's lymphoma (liver); and 5 primary hepatocellular carcinomas. CONCLUSION: There was histologic and/or immunohistochemical confirmation in 25 cases (71%). Some of the limitations in categorization of such tumors obtained by FNAB can be overcome by immunohistochemistry. Information gleaned from a precise cytologic diagnosis can sometimes only favor a particular primary site.

Serological profiles of hepatitis B carrier patients in Singapore with special reference to the frequency and significance of concurrent presence of HBsAg and anti-HBs.

Hepatitis B serological markers were investigated in 1,132 consecutive Singaporean HBV carriers. Hepatitis B surface antigen (HBsAg) and hepatitis B surface antibody (anti-HBs) were found concurrently in 234 carriers (234/1132 or 21%). Serum anti-HBs levels were more than 10 mIU/mL in 80 of these carriers (80/234 or 34%). There were no difference in HBeAg positive status, as well as HBV-DNA positive status in concurrent HBsAg/anti-HBs carriers compared to carriers without anti-HBs. Our results suggested that concurrent HBsAg and anti-HBs is a common serologic pattern in Singaporean HBV carriers.

Primary biliary cirrhosis--an uncommon disease in Singapore.

Primary biliary cirrhosis (PBC) is uncommon in Singapore. Twelve consecutive patients with PBC were seen between 1987 and 1994 at the National University Hospital. Eleven were women and the mean age at presentation was 53 years. Three patients presented with pruritus and jaundice whilst three had decompensated cirrhosis. The remaining six patients had no symptoms attributed to their liver disease when first detected, three of them presented with associated conditions including sicca syndrome and interstitial lung fibrosis, lichen planus, and carcinoma of breast. All patients had elevated serum alkaline phosphatase and positive anti-mitochondrial antibodies. Liver histology (10/12) showed Stage II disease (2), Stage III (5) and Stage IV (3). Three patients also had co-existing gall bladder stones but their endoscopic retrograde cholangiograms were normal. The mean follow-up period was 32.6 months and four patients died during follow-up. The only male patient had liver transplantation, two patients had symptomatic treatment while the rest were treated with ursodeoxycholic acid. In conclusion, local patients tended to presented relatively early in the course of the disease with 50% being asymptomatic and in the precirrhotic Stages.

A new pre-S containing recombinant hepatitis B vaccine and its effect on non-responders: a preliminary observation.

Nine healthy subjects who had no detectable antibody response when initially vaccinated with hepatitis B vaccine (plasma derived and/or yeast derived) were revaccinated with three 20 micrograms doses of a new recombinant hepatitis B (HB) vaccine (SCI-B-VAC) derived from Chinese hamster ovary (CHO) cells and consisting of both the major S protein and the minor pre-S (S1 and S2) proteins of the viral coat. Seroconversion was successful in 78% of these non-responders after the second dose and just before the third vaccination. The geometric mean titres of the antiHBs were 587 iu/l one month after the course of vaccination. This study, though preliminary, indicated that pre-S containing CHO vaccine is effective in inducing antibody response in S-vaccine non-responders and also suggested that the added pre-S region in HBV vaccine helps antiHBs production. Frequency of HLA B46 and B15 were higher in non-responders compared to normal subjects (P = 0.04) supporting the possibility that genetic factors may modulate the immune response to HB vaccination.

The effect of polyunsaturated phosphatidyl choline in the treatment of acute viral hepatitis.

BACKGROUND: Polyunsaturated phosphatidyl choline is a preparation often advocated for diseases of the liver. METHODS: In a randomized open controlled trial, a preparation of polyunsaturated phosphatidyl choline, at a dose of 900 mg orally daily, was given to 22 patients with acute viral hepatitis. A control group of 25 patients was not treated. RESULTS: Serial serum bilirubin and alanine amino transferase levels were measured up to 12 weeks. The falls in their levels after 2 and 5 weeks, and the lengths of time to their normalization, were not significantly different in the treated group compared to the control group. CONCLUSION: The results indicated that polyunsaturated phosphatidyl choline had no beneficial effect on the course of acute viral hepatitis.

Treatment of hepatitis B surface antigen carriers in the early stage of the infection using recombinant alpha-interferon with steroid priming.

BACKGROUND: Alpha-interferon has been found to inhibit hepatitis B virus (HBV) replication in Chinese patients with chronic HBV infection although a sustained effect was rarely achieved in those with normal pretreatment serum alanine amino transferase (ALT) levels. Prednisolone priming has been found to be beneficial over treatment with interferon alone in these subjects. We studied the effect of steroid pre-treatment followed by recombinant interferon alpha-2a in the treatment of asymptomatic HBV carriers with positive hepatitis Be antigen (HBeAg), hepatitis B viral DNA (HBV-DNA) and minimal changes in liver histology. METHODS: The treatment regimen included a 6-week prednisolone priming, a 2 week rest followed by 14 weeks of three times weekly 9 mega units of interferon alpha-2a injection and 52 weeks of follow-up. There were seven patients in the treatment group and seven controls. RESULTS: The mean age, pre-treatment ALT (normal in all except for one in each of the treatment and control groups), HBV-DNA levels and histological scores were similar in the two groups. Serum HBV-DNA levels fell in six patients during treatment and became undetectable in two of them by the end. During follow-up, serum HBV-DNA returned to pre-treatment levels in all patients. None of the treated patients had HBeAg sero-conversion and none of the controls had spontaneous clearance of HBV-DNA or sero-conversion of HBeAg. No improvement of liver histology was observed in any of the treated patients. There were only mild flu-like side-effects noted and interferon alpha-2a was well tolerated at the doses given among treated patients. CONCLUSION: Prednisolone priming followed by interferon alpha-2a treatment has no beneficial effect on HBV carriers in the early stages of chronic hepatitis B infection.

Bile cytology. Diagnostic role in the management of biliary obstruction.

Thirty satisfactory bile specimens from 21 patients (13 with cancer and 8 with benign conditions) were reviewed to assess the diagnostic role of bile cytology in the management of biliary obstruction. Smears were malignant in 5 instances, suspicious in 1, atypical in 2 (8 in the positive group) and negative in 22. There were no false positives. The diagnostic sensitivity was 38%, specificity 100% and overall accuracy 57%. Of interest were (1) hepatocellular carcinoma, causing obstruction in three patients; (2) metastatic colonic adenocarcinoma, causing obstruction in another instance; (3) positive cytodiagnosis in a patient who developed biliary stricture following laparoscopic cholecystectomy; and (4) detection of acidfast bacilli in bile from a case of tuberculous pseudotumor mimicking a hepatic malignancy. Bile cytology is a simple, inexpensive means of obtaining tissue confirmation of neoplastic and specific inflammatory causes of biliary obstruction. It is a useful adjunct to other techniques in this era of fiberoptic diagnostic procedures. Cell block sections should be prepared whenever possible.

Immunogenicity and safety of an inactivated hepatitis A vaccine amongst Singaporeans.

The immunogenicity and reactogenicity of an inactivated hepatitis A virus (HAV) vaccine was studied in healthy Singaporean adult volunteers. One hundred and forty healthy volunteers with normal alanine (ALT) and aspartate (AST) transaminases and no previous exposure to HAV, received three 1 ml doses (720 ELISA units) of an inactivated HAV vaccine (Smithkline Beechams Biologicals) following a 0, 1, 6 months vaccination schedule. All subjects were asked to record and grade the severity of any reactions for three consecutive days after each dose. Serum ALT and AST as well as anti-HAV were measured at 0, 1, 2, 6 and 7 months after the first vaccine dose. Anti-HAV seroconversion occurred when levels rose above 40 mIU/ml. Eighty-five percent of vaccinees seroconverted after the first innoculation and 99% after the second injection. All vaccinees seroconverted after the third dose. Geometric mean anti-HAV titers (GMTs) were, respectively, 119, 391, 4406 mIU/ml one month after each of the three doses. The most common side effect was transient pain and tenderness at the vaccination site. No elevation of ALT or AST levels were noted during the study period. The inactivated hepatitis A vaccine used in this study is safe and highly immunogenic in the local adult population. Two doses one month apart appeared to give adequate protection.

Aspiration cytology of liver abscesses. With an emphasis on diagnostic pitfalls.

Clinical and imaging features of liver abscesses are not specific. Necrotic hepatic neoplasms, primary or secondary, can mimic abscesses, and vice versa. Thirty-eight patients who had cytologic confirmed abscesses were analyzed. There was clinical, radiologic and cytologic concurrence in 27 patients. In six cases the clinically suspicious lesion turned out to be inflammatory. The remaining five were malignant. There were four amebic and three tuberculous cases in this series. Cytologically, pyogenic abscesses contained a heavy, neutrophilic, inflammatory exudate with nuclear debris. By comparison, amebic cases contained more necrotic debris, with degenerating hepatocytes and fewer inflammatory cells. Acid-fast bacilli were identified in two tuberculous abscesses; however, only one contained caseous necrotic material and epithelioid cells. A potential pitfall in the cytologic diagnosis of a case of inflammatory pseudotumor is emphasized. The diagnosis of liver abscess should be established by clinical and imaging findings in conjunction with needle aspiration.

Tuberculous pseudotumor causing biliary obstruction. Report of a case with diagnosis by fine needle aspiration biopsy and bile cytology.

Hepatobiliary tuberculosis is a rare but distinct clinical entity. We report an unusual case of biliary tract obstruction due to localized hepatic tuberculosis with periportal tuberculous adenitis. The lesion mimicked a malignancy clinically and radiologically. Fine needle aspiration biopsy revealed granulomas, epithelioid histiocytes and Langhans' giant cells. The cytodiagnosis was confirmed by identification of acid-fast bacilli in the bile cytology and isolation of Mycobacterium tuberculosis by culture. The patient responded to antituberculosis therapy. The usefulness of bile cytology in the diagnostic management of biliary tract obstruction is illustrated.

Chili--protective factor against peptic ulcer?

The aim of the present study was to determine the frequency and amount of chili taken by peptic ulcer patients and control subjects. One hundred three Chinese patients with peptic ulcer and 87 control patients were interviewed using a standard questionnaire. Those subjects who deliberately avoided chili use because of symptoms or advice from friends or medical practitioners were excluded. The median number of times of chili use per month was eight in the ulcer group (25-75% quartiles 1-30) compared to 24 (8-56) in the control group (P < 0.001). The median amount of chili used per month was 312 units (25-75% quartiles 38-899) in the ulcer group compared to 834 units (274-1892) in the control group (P < 0.001). The odds ratio of having peptic ulcer disease, adjusted for age, sex, analgesic use, and smoking by multiple logistic regression, was 0.47 (95% confidence intervals: 0.25-0.89) for subjects who had a higher intake of chili both in terms of frequency as well as amount used compared to those who took less chili. Our data support the hypothesis that chili use has a protective effect against peptic ulcer disease.

Chili protects against aspirin-induced gastroduodenal mucosal injury in humans.

Capsaicin, the pungent ingredient of chili, has a gastroprotective effect against experimental gastric mucosal injury in animals. Such an effect has not, however, been documented in humans to date. Eighteen healthy volunteers with normal index endoscopies underwent two studies four weeks apart. Each subject took 20 g chili orally with 200 ml water in one study and 200 ml water in another study. In each case this was followed half an hour later by 600 mg aspirin BP with 200 ml water. Endoscopy was repeated 6 hr later. Gastroduodenal mucosal damage was assessed by a previously validated scoring system. The median gastric injury score after chili was 1.5 compared to 4 in the control group (P < 0.05), demonstrating a gastroprotective effect of chili in human subjects.