In vitro and in vivo response of hepatocytes from hepatic nodules to the mitoinhibitory effects of phenobarbital

One of the proposed mechanisms by which phenobarbital (PB) promoteshepatocarcinogenesis in the rat is by differential mitoinhibition.However, our earlier studies indicated that PB inhibited DNAsynthesis in vitro in hepatocytes isolated from both surroundingnon-nodular liver and hepatic nodules promoted by orotic acid(OA). Since nodules generated by one promoter need not necessarilybe resistant to another promoter, the present study was undertakento determine whether foci/nodules promoted by PB itself areresistant to the mitoinhibitory effects of PB. Accordingly,rats were initiated with diethylnitrosamine (DENA, 200 mg/kgi.p) and promoted with PB (0.07% of PB as its sodium salt) intheir drinking water for 16 or 33 weeks. In vitro studies indicatedthat PB (3–5 mM) inhibited DNA synthesis induced by epidermalgrowth factor (EGF) in hepatocytes from surrounding non-nodularliver as well as from nodules promoted by PB for 33 weeks. Inanother experiment, initiated rats exposed to PB for 33 weekswere subjected to either two-thirds partial hepatectomy (PH)or sham hepatectomy. Hepatocytes were labelled with tritiatedthymidine in vivo for 48 h. Autoradiographic analysis indicatedthat in the presence of PB, the hepatocytes from both foci/nodulesand the surrounding non-nodular liver responded to PH to thesame extent. In addition, they both responded to PH less efficientlyas compared to the corresponding controls. Further, initiatedrats exposed to PB for 16 weeks when subjected to PH and killed4 weeks thereafter, the percentage area occupied by     

Perturbations of endogenous levels of orotic acid and carcinogenesis: effect of an arginine-deficient diet and carbamyl aspartate on hepatocarcinogenesis in the rat and the mouse

Feeding excess orotic acid (OA) In the diet promotes the carcinogenicprocess in different organs Including the liver. A number ofmetabolic and genetic disorders are associated with Increasedsynthesis of endogenous OA and some of these disorders appearto pose an Increased risk of liver cancer development. Thisstudy therefore examines whether excess OA of endogenous originalso exerts a promoting effect on hepatocarcinogenesis in themouse and the rat. Increased endogenous synthesis of OA wasachieved by (i) feeding a diet deficient in arginine (AD) and(ii) feedIng excess dietary carbamylaspartate (CA), a precursorfor the synthesis of OA. A single dose of diethylnitrosamine(DENA) was given i.p. to male Fischer 344 rats (200 mg/kg) orto male DBA/2 mice (90 mg/kg). One week later they were placedon either AD diet or the same diet supplemented with 1.3% arginine(AS) for a total of weeks. Two-thirds partial hepatectomy (PH)was performed at the end of the second week. All animals werethen transferred to a control semisynthetic basal diet for atotal of 20 weeks before they were killed. The results indicatedthat AD diet increased the incidence of hepatic nodules in bothrats (percentage area occupied by nodules was 4.7 ± 0.4in the AD group compared to a control value of 0.7 ±0.5) and mice (4/10 mice had nodules >5 mm diameter in theAD group while none in the AS group had such large nodules).In another experiment male Fischer 344 rats similarly initiatedwIth DENA were exposed to either basal diet or basal diet containing2% CA for 4 weeks coupled with PH performed at the end of thesecond week. This regimen was followed by 20 weeks of feedingbasal diet to both groups. Rats given CA developed larger hepatlcfoci and nodules (0.84 ± 0.56 mm³) compared to the controlgroup, which was fed basal diet throughout the experiment (0.07± 0.03 mm³) Further, both AD diet and dietary CA, likedietary OA, induced an increase In hepatic uridine nucleotides.Taken together, these results suggest that Increased levelsof endogenously synthesized OA, like exo genously supplied excessOA, can induce an Imbalance in hepatic nucleotide pools andcan exert a promoting effect on hepatocarcinogenesis.     

Manual small incision cataract surgery: a viable option for cataract with pseudoexfoliation

International Ophthalmology - The aim of the study was to assess the spectrum of clinical presentation, intraoperative challenges and immediate surgical outcome of cataract patients with...     

Severity of diabetic retinopathy and its relationship with age at onset of diabetes mellitus in India: A multicentric study

Purpose:To present clinical profile and risk factors of sight-threatening diabetic retinopathy (STDR) among people with age of onset of diabetes (AOD) <25 versus ≥25 years.Methods:A retrospective chart analysis of consecutive patients with diabetic retinopathy (DR) n = 654) treated at 14 eye care centers across India between 2018 and 2019 was performed. Patients were divided into two groups, Group 1: AOD <25 years and Group 2: AOD ≥25 years. DR and diabetic macular edema (DME) were classified using the International Clinical Classification of DR severity scale. STDR included severe nonproliferative DR (NPDR), proliferative DR (PDR), and moderate to severe DME. A multilevel mixed-effects model was used for comparison between two groups: 1) Patients with DR and AOD <25 years and 2) Patients with DR and AOD ≥25 years. Bivariate and multivariate regression analyses were used to evaluate risk factors between the two groups.Results:A total of 654 patients were included, 161 (307 eyes) in AOD <25 and 493 (927 eyes) in AOD >25 group. There was a higher prevalence of PDR with high-risk characteristics in AOD <25 group (24% vs. 12%) at baseline and 12-month follow-up (25% vs. 6%); P < 0.001. Systolic hypertension and poor glycemic control were risk factors in both groups, with no difference in these modifiable risk factors between groups.Conclusion:People with youth-onset DM are likely to present with severer form of STDR despite similar modifiable risk factors. Therefore, strict control of systolic blood pressure, glycemic status, and regular screening for DR are recommended to reduce the risk of STDR irrespective of the age of onset of diabetes.     

Susceptibility of dimethylnitrosamine induced O-methylguanine containing regions in in vivo replicated, hybrid rat liver DNA towards S1 nuclease

Experiments were designed to characterize the carcinogen-modified rat liver DNA following in vivo replication. Replicated hybrid DNA of density 1.718 g/cm³ and greater, containing [¹⁴C]dimethylnitrosamine-induced methylated products and 5-bromo-2-deoxy-uridine was isolated and treated with S₁ nuclease. The extent of destabilized regions in such a hybrid DNA was determined by monitoring the release by S₁ nuclease of O⁶-methylguanine, a lesion known to cause miscoding and N-7-methylguanine, a lesion that apparently does not miscode. The results indicate that when such replicated rat liver hybrid DNA is treated with S₁ nuclease, O⁶-methylguanine containing regions compared to those containing N-7-methylguanine are preferentially digested by the enzyme.     

5-Azacytidine potentiates initiation induced by carcinogens in rat liver

To test the validity of the hypothesis that hypomethylationof DNA plays an important role in the initiation of carcinogenicprocess, 5-azacytidine (5-AzC) (10 mg/kg), an inhibitor of DNAmethylation, was given to rats during the phase of repair synthesisinduced by the three carcinogens, benzo[a]-pyrene (200 mg/kg),N-methyl-N-nltrosourea (60 mg/kg) and 1,2-dimethylhydrazine(1,2-DMH) (100 mg/kg). The initiated hepatocytes in the liverwere assayed as the -glutamyltransferase (-GT) positive fociformed following a 2-week selection regimen consisting of dietary0.02% 2-acetylaminofluorene coupled with a necrogenic dose ofCCl₄. The results obtained indicate that with all three carcinogens,administration of 5-AzC during repair synthesis increased theincidence of initiated hepatocytes, for example 10–20foci/cm² in 5-AzC and carcinogen-treated rats compared with3–5 foci/cm² in rats treated with carcinogen only. Administrationof [³H]-5-azadeoxycytidine during the repair synthesis inducedby 1,2-DMH further showed that 0.019 mol% of cytosine residuesin DNA were substituted by the analogue, indicating that incorporationof 5-AzC occurs during repair synthesis. In the absence of thecarcinogen, 5-AzC given after a two thirds partial hepatectomy,when its incorporation should be maximum, failed to induce any-GT positive foci. The results suggest that hypomethylationof DNA per se may not be sufficient for initiation. Perhapstwo events might be necessary for initiation, the first causedby the carcinogen and a second involving hypomethylation ofDNA.     

A comparative study of nonpharmacological methods to reduce pain in neonates

A randomized study was done to compare non pharmacological methods to reduce the pain of heel pricks in 104 stable term neonates. Non-nutritive sucking (NNS), rocking, massage, sucrose (20 percent), distilled water (DW) and expressed breast milk (EBM) were used as pain reducing agents. Duration of cry and Douleur Aigu? du Nouveau né (DAN) score were used to assess pain. Physiological parameters were also recorded before and after the stimulus. At 30 seconds after the stimulus, the pain scores were lowest in the sucrose group but this was not sustained at 1, 2 and 4 minutes. At 2 and 4 minutes pain scores were lowest in the NNS and rocking groups as compared to sucrose, distilled water, expressed breast milk and massage. The total duration of crying was also lowest in the NNS and rocking groups. Physiological parameters were comparable in all groups. Babies who were in Prechtl State 1 and 2 (sleeping) at the time of stimulus showed significantly lesser response to pain compared to babies who were awake. This was seen in all the intervention groups. In conclusion, our study suggests that rocking or giving a baby a pacifier are more effective non-pharmacological analgesics than EBM, DW, sucrose or massage for the pain of heel pricks in neonates. A calm or sleeping state before a painful procedure also appears to decrease crying and pain scores.     

Tumor microenvironment modulates hyaluronan expression: the lactate effect

Hyaluronan (HA) synthesis is a tightly regulated process and is partly controlled by the microenvironment (e.g., lactate concentration). Experimental evidence has indicated that the melanoma cells that synthesize large amounts of HA exhibit enhanced tumor cell growth and increased metastatic capacity compared to those expressing smaller amounts. Because most studies have examined HA expression on melanoma cells in vitro, we compared the patterns of HA expression by B16-F1 and B16-F10 melanoma cells in vitro and in situ. Cell surface HA expression was assessed with the HA-binding peptide Pep-1. B16-F1 melanoma cells showed significantly higher levels of Pep-1 binding compared with B16-F10 cells in vitro. On the other hand, expression levels of HA were comparable between B16-F1 and B16-F10 melanoma cells in cryostat sections. These results show that B16-F1 cells express high levels of HA in vitro and in vivo, while B16-F10 cells express high concentrations of HA only in the context of skin tumors. Finally, B16-F10 melanoma cells, but not B16-F1 cells, expressed high concentrations of HA after stimulation with lactate. We propose that components of the tumor microenvironment (e.g., lactate) can induce melanoma cells to express HA and thus acquire an aggressive phenotype.