The role of coronary calcium score in the risk assessment of liver transplant candidates.

BACKGROUND: Coronary artery disease (CAD) is a common cause of morbidity and mortality in liver transplant (LT) recipients. To date there is no consensus on the preferred screening tests to detect CAD in the pre-LT population. Therefore the aim of this study was to: 1) evaluate the utility of a noninvasive tool (cardiac computerized tomography [CT] scan); and 2) determine the prevalence of CAD in low-risk LT candidates. METHODS: Using our transplant database we identified all LT candidates classified as low risk for CAD. All low-risk candidates underwent cardiac CT scan for coronary calcium score (CCS) estimation. Those with CCS >100 underwent coronary angiogram, and those with <100 underwent stress test and if stress test was positive then coronary angiography was performed. The Agatston calcium score was classified as: normal (0), mild (1-100), moderate (101-400), severe (401-1,000), or extensive (>1,000). RESULTS: Eighty-five LT candidates were classified as low risk and underwent cardiac CT scan. The mean calcium score was 325 (range, 0-3,707). In our study cohort, 21% had normal CCS score, 43% mild, 13% moderate, 11% severe, and 12% extensive. A calcium score >400 was significantly associated with CAD on angiography (P = .02). Although male sex was significantly associated with the presence of CAD (P = .006), there was no correlation with age, ethnicity, liver diagnosis, or Model for End-Stage Liver Disease score. CONCLUSIONS: Prevalence of asymptomatic CAD in this low-risk population is relatively high. Cardiac CT is well tolerated and is a useful noninvasive screening tool in LT candidates. Future studies to determine its utility as a prognostic tool after LT will be invaluable.

Anti-tumorigenic action of 2-[piperidinoethoxyphenyl]-3-[4-hydroxyphenyl]-2H-benzo(b)pyran: evidence for involvement of GPR30/EGFR signaling pathway.

OBJECTIVE: The aim of the present study was to investigate the effect of non-steroidal, pure antiestrogenic benzopyran derivative i.e., 2-[piperidinoethoxyphenyl]-3-[4-hydroxyphenyl]-2H-benzo(b)pyran (K-1) on the growth of human endometrial cancer cells in vivo and in vitro and to elucidate its mechanism of action. METHODS: Cell proliferation was assayed by measuring the incorporation of 5'-bromo-2'-deoxyuridine in Ishikawa and primary endometrial cancer cells. The expression of proliferation and apoptotic markers was analyzed by immunoblotting. The effect of K-1 on GPR30-regulated proteins was analyzed by ELISA and by immunoblotting. Nude mice bearing subcutaneous implanted-Ishikawa tumors, were treated for 14days with K-1 (200µg/kg body weight/day/orally). The proliferation markers, GPR30-regulated proteins and apoptotic markers were analyzed by immunoblotting in tumor xenograft. The apoptotic effect of compound K-1 was determined by TUNEL assay. RESULTS: Compound K-1 inhibited proliferation of endometrial adenocarcinoma cells and decreased the expression of proliferation markers. It caused apoptosis by increasing the expression of apoptotic markers (NOXA, PUMAα) and reducing the expression of p-CREB and BclxL. Compound interfered with GPR30-regulated-EGFR activation, decreased p-ERK, p-c-jun, c-fos, cyclinD1 and c-myc expression. Treatment of tumor-bearing mice with K-1 resulted in a significant decrease in tumor volume and weight. Decreased expression of p-ERK and its downstream molecules and increased expression of apoptotic markers were observed in tumor in K-1 treated animals. CONCLUSION: Findings suggest the potent inhibitory effect of compound K-1 on endometrial cancer cellular growth (in-vitro) and on tumor size (in-vivo) which is mediated at least, in part, by interference with GPR30-signaling.

Peptidyl prolyl isomerase, Pin1 is a potential target for enhancing the therapeutic efficacy of etoposide.

The peptidyl prolyl isomerase (Pin1) that induces cis-trans isomerization of the peptide bond involving serine/threonine-proline has recently been shown to regulate the activity of many phosphoproteins including the ones involved in damage response pathways. We investigated Pin1 as a potential target for enhancing the efficacy of anticancer therapy by studying the effects of juglone, a Pin1 inhibitor on the cytotoxicity of etoposide (a widely used anticancer drug that targets topoisomerase IIα) in human tumor cell lines. Treatment of cells with juglone synergistically enhanced the cytotoxicity of etoposide (loss of clonogenicity) with a tenfold increase when etoposide treatment preceded juglone exposure. On the other hand, the toxicity was than additive when the treatment protocol was reversed (i.e exposure to juglone followed by etoposide treatment). This suggests that Pin1 inhibition possibly reduces the induction of initial DNA damage by etoposide, which was supported by a decrease in the levels of chromatin bound topoIIα. Increase in the etoposide induced toxicity by juglone appeared to be mainly due to enhanced mitotic cell death linked to cytogenetic damage, although a moderate increase in interphase (apoptotic) death was also evident as revealed by DNA degradation (hypodiploid population and TUNEL assay). Since the level of Pin1 is found to be higher in cancer cells, this enzyme could be a potential target for developing an adjuvant to enhance the efficacy of anticancer therapies.

Detection of influenza virus induced ultrastructural changes and DNA damage.

The influenza virus generally causes damage to epithelial cells of respiratory tract and infection of cells with this virus often results in cell death with apoptotic characteristics. Reports are available implicating influenza virus as a causative agent of chromosomal aberrations in cells and culture. The objective of this study was to analyze the process of cell death caused by influenza virus (A/Udorn/317/72, H3N2) infection in cultured HeLa cells by electron microscopy and comet assay. The apoptotic study was performed using light microscopy electron microscopy and comet assay to observe the changes in cell morphology and DNA fragmentation. HeLa cells, infected with influenza virus were harvested at various time periods to observe the ultrastructural changes. This infection gave rise to nuclear fragmentation and chromatin condensation accompanied by chromosomal DNA fragmentation into oligonucleosomes. The pattern of comet assay revealed that the apoptosis occurred due to fragmentation of the DNA of the cells which reached the maximum level at 36 h post infection. Ultrastructural study showed extensive chromatin condensation and nuclear fragmentation which are the characteristic features of apoptosis.

Radiation responses of Sf9, a highly radioresistant lepidopteran insect cell line.

PURPOSE: Lepidopteran insect cells are known to exhibit very high radioresistance. Although very effective DNA excision-repair has been proposed as a contributing factor, a detailed understanding of insect cell radiation responses has not yet been obtained. Therefore, the study was carried out to understand the in vitro radiation responses of Sf9 lepidopteran cells. MATERIALS AND METHODS: Exponentially growing asynchronous Sf9 cells (derived from ovaries of Spodoptera frugiperda) were exposed to gamma-radiation doses of 2-200 Gy. Cell survival, growth inhibition, cell cycle progression delay, alterations in cell morphology as well as induction of DNA damage, micronuclei and apoptosis were studied at various post-irradiation time intervals. RESULTS: Biphasic survival response curves were obtained with D0 rising from 20 Gy (at doses < or = 60 Gy) to 85 Gy (between 60 and 200 Gy), corroborating earlier reports on lepidopteran cells. An additional downward deviation at 2 Gy indicated a hypersensitive response. Dose-dependent growth inhibition with a transient G2 delay starting 12 h and extending up to 48-96 h was observed at doses of 10-200 Gy, while a brief G1/S transition delay was observed only at higher doses (> or = 100 Gy). Significant DNA damage was detected only at 20 Gy and higher doses, in contrast with human cells that showed similar damage at 2 Gy. Interestingly, micronuclei were not induced at any of the doses tested, although spontaneous micronucleation was evident in <1% of cells. Lack of micronucleus induction even at doses that induced significant DNA damage and a transient G2 block (20-50 Gy) strongly indicated a role of holocentric lepidopteran chromosomes. Apoptosis was detected only in a small proportion of cells (3%) exposed to 200 Gy, and cell/nucleus size and granularity increased by 72-96 h post-irradiation in a dose-dependent manner. Sf9 nucleoids extracted at 2 M NaCl showed higher compactness than the nucleoids prepared from human cells. CONCLUSIONS: It is clearly shown that lepidopteran cells are highly resistant to the induction of DNA damage and micronuclei, and display very low induction of apoptosis at doses up to 200 Gy. While the lack of micronucleus induction seems to be primarily due to the holocentric nature of their chromosomes, certain unique signalling pathways might be responsible for the low induction of apoptosis. Factors causing protection of Sf9 cellular DNA from radiation-induced damage are presently being investigated.

Relation between plasma leptin and anthropometric and metabolic covariates in lean and obese diabetic and hyperlipidaemic Asian Northern Indian subjects.

This study investigated the relationship of plasma leptin to obesity, diabetes and hyperlipidaemia in Asian Northern Indian subjects, considered to have a predisposition to abdominal obesity and metabolic syndrome. A total of 72 subjects, subcategorised into lean and obese healthy subjects, lean and obese Type 2 diabetic and lean and obese non-diabetic hyperlipidaemic subjects were recruited. High leptin values were observed in all obese groups, and obese diabetic patients showed the highest levels. In lean and obese diabetic subjects, plasma leptin did not show any correlation to any index of glycaemia. When all lean and all obese subjects were analysed in two separate groups, body mass index (BMI), percent total body fat, and body density significantly correlated with the plasma leptin levels (p<0.05). Leptin values, when correlated to all variables in all patients taken together, showed the greatest magnitude of correlation with BMI (r=0.64), percent total body fat (r=0.67), and waist circumference (r=0.51). Strong inverse correlation was seen with body density (r=-0.67). Levels of serum insulin did not show any correlation with leptin levels in all subjects combined, and separately in various groups. Multiple linear regression analysis performed in obese, non-diabetic and normolipidaemic subjects, all Type 2 diabetic and all non-diabetic hyperlipidaemic subjects separately showed that percent total body fat is the only significant predictor of plasma leptin concentration in all the 3 groups. The present study suggests that plasma leptin has a strong positive correlation with percent total body fat in Asian Northern Indian subjects. Among other components of metabolic syndrome, only abdominal obesity is weakly correlated to serum leptin levels.

What is hypertension in chronic haemodialysis? The role of interdialytic blood pressure monitoring.

BACKGROUND: Hypertension in chronic haemodialysis patients contributes significantly to morbidity and mortality. Treatment decisions are usually based on predialysis readings, which may not accurately reflect control during the interdialytic period. METHODS: We studied 40 randomly selected subjects on haemodialysis and compared readings by different methods at set times during the dialysis session with the 48-h interdialytic ambulatory readings. Conventional sphygmomanometer, automated Dinamap and Tm 2421(A&D) ambulatory monitor were used for BP measurements. RESULTS: Conventional sphygmomanometry and self measured automatic readings (Dinamap) were highly correlated (systolic r=0.93, P<0.001; diastolic r=0.90, P<0.001). Mean blood pressure on arrival ((PreC(0)) 158 mmHg systolic, 80 mmHg diastolic and 106 mmHg mean) significantly overestimated the mean ambulatory reading during the 6 h prior to attendance ((preAm(6h)) systolic 147 (P<0.01), diastolic 75 (P<0.01), mean 99 (P<0.01)). Fifteen patients (41%) demonstrated a marked difference (>20/10 mmHg) between the PreC(0) and preAm(6h) (white-coat effect) persisting in seven patients (19%) after a period of rest 10 min predialysis (preC(10)) and present even in self-recorded Dinamap readings. There was a significant negative relationship between the systolic rise and the number of months on dialysis (P<0.05). Mean ambulatory BP on interdialytic day 2 was significantly greater than on day 1 whereas the awake-sleep differences were less on day 2 than day 1, both perhaps reflecting differences in volume status. The 20 min post-dialysis measurement (PoC(20)) for systolic, diastolic, and mean, unlike predialysis (PreC(0) and preC(10)), onset (onC) and end of dialysis readings (enC) did not differ significantly from 48 h interdialytic means. CONCLUSIONS: The best representation of interdialytic pressure was the 20-min post-dialysis reading. Walk-in predialysis pressures overestimate mean interdialytic pressures due to a high incidence of white-coat effect, which shows some habituation with time on dialysis. Ambulatory monitoring has a role in evaluating persistent poor blood pressure control in haemodialysis patients.

Is there a rationale for rationing chronic dialysis? A hospital based cohort study of factors affecting survival and morbidity.

OBJECTIVES: To determine factors influencing survival and need for hospitalisation in patients needing dialysis, and to define the potential basis for rationing access to renal replacement therapy. DESIGN: Hospital based cohort study of all patients starting dialysis over a 4 year recruitment period (follow up 15-63 months). Groups were defined on the basis of age, comorbidity, functional status, and whether dialysis initiation was planned or unplanned. SETTING: Renal unit in a district general hospital, which acts as the main renal referral centre for four other such hospitals and serves a population of about 1.15 million people. SUBJECTS: 292 patients, mean age 61.3 years (18-92 years, SD 15.8), of whom 193 (66%) were male, and 59 (20%) were patients with diabetes. Dialysis initiation was planned in 163 (56%) patients and unplanned in 129 (44%). MAIN OUTCOME MEASURES: Overall survival, 1 year survival, and hospitalisation rate. RESULTS: Factors affecting survival in the Cox's proportional hazard model were Karnofsky performance score at presentation (hazard ratio 0.979, 95% confidence interval 0.972 to 0. 986), comorbidity severity score (1.240, 1.131 to 1.340), age (1.036, 1.018 to 1.054), and myeloma (2.15, 1.140 to 4.042). The Karnofsky performance score used 3 months before presentation was significant (0.970, 0.956 to 0.981), as was unplanned presentation in this model (1.796, 1.233 to 2.617). Using these factors, a high risk group of 26 patients was defined, with 19.2% 1 year survival. Denying dialysis to this group would save 3.2% of the total cost of the chronic programme but would sacrifice five long term survivors. Less rigorous definition of the high risk group would save more money but lose more long term survivors. CONCLUSIONS: Severity of comorbid conditions and functional capacity are more important than age in predicting survival and morbidity of patients on dialysis. Late referral for dialysis affects survival adversely. Denial of dialysis to patients in an extremely high risk group, defined by a new stratification based on logistic regression, would be of debatable benefit.

Radioprotective effects of DNA ligands Hoechst-33342 and 33258 in whole body irradiated mice.

Radioprotective effects of bisbenzimidole derived DNA ligands Hoechst-33342 (H-342) and Hoechst-33258 (H-258) have been investigated in whole body irradiated stain-A and Balb/c mice (Co-60 Gamma-ray, absorbed doses of 2.5 to 10 Gy delivered at dose rates of 0.01 to 0.50 Gy/min). Biodistribution of Hoechst dyes (2 or 5 mg/kg, body wt., i.v.) and their effects on cell cycle kinetics in bone marrow were studied by flow cytometry. Protection against radiation-induced chromosomal aberrations, micronuclei formation, alterations in DNA content dispersion, inhibition of erythropoiesis and animal lethality were investigated. Significant amount of DNA bound Hoechst could be observed in liver, intestine, kidney and brain for more than 14 days after its administration, while in the bone marrow cells, a reduction in the bound Hoechst was noticed after 7 days. H-342 significantly reduced the radiation-induced chromosome aberrations mainly due to a decrease in the frequency of acentrics (nearly 30%), while a marginal decrease (10%) in the dicentrics was observed at all the dose rates studied. Both H-342 and H-258 reduced the radiation-induced micronuclei formation in a dose dependent manner (2-10 mg/kg body wt.) and this protective effect was observed up to 6 days after the administration. Neither of the two compounds induced any cytogenetic damage in the bone marrow cells of unirradiated animals nor induced tumours at the doses used here (< 5 mg/kg, body wt. i.v.). Reduction in cytogenetic damage of bone marrow cells led to a faster recovery of erythropoesis as observed by increased PCE/NCE ratio in the peripheral blood erythrocytes of the animals which received Hoechst before irradiation. H-258 (5 mg/kg body wt.) given 18 hr before irradiation reduced radiation-induced animal death (5-9 Gy), while no significant effect was observed at higher doses (10 Gy). However, H-342, which has a higher cell permeability, even at a lower dose (2 mg/kg body wt.) showed significant protection at 10 Gy. The protective effects could be enhanced further, by combining these DNA binding agents with the glucose analogue, 2-deoxy-D-glucose (2-DG) which has been shown earlier to protect bone marrow cells against radiation damage.

Low serum vitamin B12 levels in chronic high-flux haemodialysis patients.

The occurrence of vitamin B12 (B12) deficiency in chronic haemodialysis patients and the need for its supplementation in these patients are still matters of debate. We measured serial predialysis serum B12 levels, at 3- to 6-month intervals, in 67 unselected patients on our high-flux haemodialysis programme. Over a 12-month period, there was a significant fall in serum B12 from 497 +/- 200 (SD) to 391 +/- 131 ng/l (p < 0.001). 22 patients developed subnormal serum B12 levels and were commenced on hydroxocobalamin supplements. We were unable to demonstrate B12 clearance during dialysis using blood side studies. Measurement of B12 in the dialysate showed that 0-4.5 microg B12 was cleared per dialysis. Using these B12 measurements, in vivo B12 clearance was estimated at 9.1 ml/min. Dietary studies on 24 unselected patients showed borderline or low B12 intake in 4 patients. Absorption studies by whole-body counting on 6 patients using 57Co and 58Co showed normal B12 absorption. The same radioisotope studies demonstrated no B12 adsorption to the dialyser membrane. This study demonstrates that low serum B12 levels occur in high-flux haemodialysis patients and that losses during dialysis and dietary deficiency may be contributing factors.

The Whitaker hook in the treatment of posterior urethral valves.

OBJECTIVE: To assess the efficacy of the Whitaker diathermy hook in the treatment of posterior urethral valves (PUV). PATIENTS AND METHODS: Seventeen patients with PUV underwent primary treatment under general anaesthesia using a diathermy hook. The results were assessed by the clinical course of the patient, serial measurements of serum creatinine level and repeat cystography 6 weeks and 6 months after treatment. RESULTS: There were no complications in 12 patients and they required no further treatment of the valves. Three patients required a repeat procedure which resolved the urethral obstruction. Two patients continued to show incomplete disruption of the valves and required endoscopic treatment. Two patients developed ureteric obstruction after disruption of the valves, which was treated by supravesical diversion and later reversed with no further treatment of the valves. CONCLUSIONS: The Whitaker diathermy hook is a safe and effective instrument which has not gained its rightful place in the treatment of patients with PUV.

Presence of embryotoxic factor in the sera of neonates affected by myelomeningocele: a study on chick embryo.

An experimental study on chick embryos was conducted where 0.02 mL of serum, taken from five neonates (1 to 7 days of age) with myelomeningocele was injected separately into the fertile white leghorn eggs, at zero hour of incubation. A total of 150 experimental and 135 control eggs were injected. It was observed that the embryos of chick exposed to experimental serum had an increased frequency of embryonic death and/or gross abnormalities as compared with the control group (chi 2 = 32.07; P less than .001). The anomalies observed were those of neural tube, gastrointestinal system, and musculoskeletal system. In addition, there was a generalized growth retardation among experimental embryos. These malformations are probably due to an embryotoxic factor, which is present in the serum of the baby with myelomeningocele and has been transmitted transplacentally.

Accessory limb associated with spinal bifida.

A neonate with three legs associated with spina bifida cystica and accessory rudimentary external genitalia is presented. A detailed radiologic workup was done preoperatively. The nomenclature and morphogenesis of this anomaly is briefly discussed.

Hemangiolymphangioma of the urinary bladder in a child.

A rare case of hemangiolymphangioma of the urinary bladder with associated cutaneous hemangiomas of the shaft and glans penis and the scrotum is presented. Unusual features of clinical presentation, noninvasive diagnosis, and the surgical management are discussed.