Effect of hepatitis B virus on steatosis in hepatitis C virus co-infected subjects: A multi-centre study and systematic review.

It remains unclear whether hepatitis B virus (HBV) infection may modify the severity of viral steatosis in patients coinfected with chronic hepatitis C virus (HCV). We examined the influence of coinfection with HBV on prevalence of steatosis in chronic hepatitis C in a multi-centre cohort of HBV-HCV subjects, and by performing a systematic review and meta-analysis of the literature. We centrally and blindly assessed steatosis prevalence and severity in a cohort of HBV-HCV coinfected subjects compared to HCV and HBV monoinfected controls and we performed a systematic review of studies addressing the prevalence of steatosis in HBV-HCV subjects compared to HCV controls. In the clinical cohort, we included 85 HBV-HCV, 69 HBV and 112 HCV subjects from 16 international centres. There was no significant difference in steatosis prevalence between the HBV-HCV and the HCV groups (33% vs 45%, P = .11). In subgroup analysis, lean HBV-HCV subjects with detectable HBV DNA had less steatosis than lean HCV subjects matched for HCV viremia (15% vs 45%, P = .02). Our literature search identified 5 additional studies included in a systematic review. Overall, prevalence of steatosis > 5% was similar in HBV-HCV infection compared to HCV (pooled odds ratio [OR] 0.91, 95% CI 0.53-1.6) although there was significant heterogeneity (I2 69%, P = .007). In conclusion, although the prevalence of steatosis is similar in HBV-HCV compared to HCV subjects, our analysis suggests that there may be an inhibitory effect of HCV-induced steatogenesis by HBV in certain subgroups of patients.

Inosine triphosphatase deficiency helps predict anaemia, anaemia management and response in chronic hepatitis C therapy.

Anaemia frequently complicates peginterferon/ribavirin therapy for chronic hepatitis C infection. Better prediction of anaemia, ribavirin dose reduction or erythropoietin (EPO) need, may enhance patient management. Inosine triphosphatase (ITPA) genetic variants are associated with ribavirin-induced anaemia and dose reduction; however, their impact in real-life clinic patient cohorts remains to be defined. We studied 193 clinic patients with chronic hepatitis C infection of mixed viral genotype (genotype 1/4 n = 123, genotype 2/3, n = 70) treated with peginterferon/ribavirin. Patients were genotyped for ITPA polymorphisms rs1127354 and rs7270101 using Taqman primers. Hardy-Weinberg equilibrium was present. Estimated ITPA deficiency was graded on severity (0-3, no deficiency/mild/moderate/severe, n = 126/40/24/3, respectively). Multivariable models tested the association with anaemia at 4 weeks of treatment [including decline in haemoglobin (g/dL); haemoglobin <10 g/dL and haemoglobin decline >3 g/dL]; ribavirin dose reduction and EPO use and explored sustained viral response (SVR) to peginterferon/ribavirin. More severe ITPA deficiency was associated with less reduction in haemoglobin level (P <0.001; R(2) = 0.34), less ribavirin dose reduction (OR 0.42; (95% CI = 0.23-0.77); P = 0.005) and less EPO use [OR 0.53; (0.30-0.94); P = 0.029]. ITPA deficiency was associated with SVR [OR: 1.70; (1.02-2.83); P = 0.041] independently of clinical covariates (adjusted R(2) = 0.31). In this clinical cohort, ITPA deficiency helped predict the risk of on-treatment anaemia, ribavirin dose reduction, need for EPO support and was associated with SVR. For patients on HCV regimens including peginterferon/ribavirin, testing for ITPA deficiency may have clinical utility.

DNA repair following ultraviolet and N-ethyl-N-nitrosourea treatment of cells cultured from human fetal brain, intestine, kidney, liver, and skin.

DNA excision repair was measured in cell cultures derived from human fetal brain, intestine, kidney, liver, and skin following ultraviolet (UV) irradiation and N-ethyl-N-nitrosourea (ENU) treatment. Cells in early passages were exposed to 5 or 10 J of UV radiation per sq m or to 25 microM to 3.5 mM ENU. DNA excision repair was determined by (a) scintillation counting and autoradiography to measure unscheduled DNA synthesis (UDS) and (b) the UV-endonuclease-sensitive site assay to measure pyrimidine dimers directly. The level of UDS following treatment of these cell cultures with UV was both time and dose dependent. UDS also increased with increasing doses of ENU up to 350 microM but decreased at doses greater than 500 microM. Cells derived from human fetal brain, kidney, and liver appeared to exhibit lower (50 to 80%) levels of UDS following UV irradiation or ENU treatment than did cells cultured from human fetal skin or intestine. The loss of UV-endonuclease-sensitive sites assayed in skin, liver, and kidney cells over a 24-hr period confirmed the differences observed by UDS in these cells. Skin cells removed 50% of the initial pyrimidine dimers from their DNA within an 8-hr period and 65 to 86% in 24 hr. Kidney and liver cells, on the other hand, removed only 28 and 32% of the initial dimers, respectively, over a 24-hr period. The data suggest differential excision repair responses following UV irradiation and ENU treatment of cells derived from different human fetal organs.

Growth-promoting effect of 12-O-tetradecanoylphorbol-13-acetate on cultured normal human fetal kidney epithelial cells.

Normal human epithelial cells have been reported to be sensitive to growth inhibition by 12-O-tetradecanoylphorbol-13-acetate in contrast to neoplastic counterparts. Our studies with normal human fetal kidney epithelial cells, however, show that 12-O-tetradecanoylphorbol-13-acetate may increase cell density and promote the growth of these cells at early passages. The result, together with three previous reports showing similar effects in normal human melanocytes, prostatic epithelial cells, and an unidentified cell type in human epidermal cell culture, indicates that human cells may exhibit divergent responses to 12-O-tetradecanoyl-phorbol-13-acetate for induction of cellular differentiation or proliferation.

Contact insensitivity of a subpopulation of normal human fetal kidney epithelial cells and of human carcinoma cell lines.

Early passage normal human fetal kidney epithelial cells were inoculated on top of a confluent monolayer of X-ray lethally irradiated human fibroblasts to determine the colony-forming ability of these epithelial cells. The results indicate that the great majority of the epithelial cells did not have the clonogenic ability on the fibroblast cell mat, although they were capable of colony formation on plastic surface without the cell mat. A small subpopulation of these epithelial cells, however, was able to proliferate on the cell mat. These contact-insensitive fetal epithelial cells were found to be deficient in gap junction-mediated intercellular communication, to contain keratin and gamma-glutamyl transpeptidase but not fibronectin. These contact-insensitive cells appear to have greater proliferative potential than the parental cell population and to exist transiently in early passage but not in late passage culture. The ability of proliferation on cell mat was found to be shared by 22 different human carcinoma cell lines that were tested. This unique clonogenic ability of normal contact-insensitive and human carcinoma cells on the cell mat could provide a selection method for presumptive normal stem and tumor cells and for an assay for screening potential antitumor drugs and assessing the efficacy of chemotherapeutic drugs against a given tumor.

Impaired K(+) homeostasis and altered electrophysiological properties of post-traumatic hippocampal glia.

Traumatic brain injury (TBI) can be associated with memory impairment, cognitive deficits, or seizures, all of which can reflect altered hippocampal function. Whereas previous studies have focused on the involvement of neuronal loss in post-traumatic hippocampus, there has been relatively little understanding of changes in ionic homeostasis, failure of which can result in neuronal hyperexcitability and abnormal synchronization. Because glia play a crucial role in the homeostasis of the brain microenvironment, we investigated the effects of TBI on rat hippocampal glia. Using a fluid percussion injury (FPI) model and patch-clamp recordings from hippocampal slices, we have found impaired glial physiology 2 d after FPI. Electrophysiologically, we observed reduction in transient outward and inward K(+) currents. To assess the functional consequences of these glial changes, field potentials and extracellular K(+) activity were recorded in area CA3 during antidromic stimulation. An abnormal extracellular K(+) accumulation was observed in the post-traumatic hippocampal slices, accompanied by the appearance of CA3 afterdischarges. After pharmacological blockade of excitatory synapses and of K(+) inward currents, uninjured slices showed the same altered K(+) accumulation in the absence of abnormal neuronal activity. We suggest that TBI causes loss of K(+) conductance in hippocampal glia that results in the failure of glial K(+) homeostasis, which in turn promotes abnormal neuronal function. These findings provide a new potential mechanistic link between traumatic brain injury and subsequent development of disorders such as memory loss, cognitive decline, seizures, and epilepsy.

[Hepatic haemorrhage in pregnancy: a case report]. / Emorragia epatica in gravidanza: caso clinico.

Spontaneous hepatic haemorrhage in pregnancy (SHHP) is a rare event (1 woman out of 15,000). It is generally considered as an advanced state of the microangiopathic hemolytic anemia (HELLP, Hemolysis, Elevated Liver enzyme levels, Low Platelet count). Furthermore, the HELLP is considered as a different form of preeclampsia. The patient, a 33-year-old-woman at 30 weeks' gestation, was admitted to hospital for preeclampsia, underwent an emergency Stark caesarean section with the extraction of an alive foetus and evidence of massive intraperitonal haemorrhage from a large hepatic haematoma. A haemostasis with gauzes of Surgicel was performed, with consequent arrest of the haemorrhage. After approximately 6 hours, a recurrence of the intraperitonal haemorrhage led to a new surgical intervention with hepatic packing with gauzes. After 4 days the patient died. The etiopathogenesis of disease is uncertain, both foetal and maternal mortality are high, and the slight number of reported cases (27) of SPPH from HELLP in international literature offer elements for debate. The following points have been put forward: 1. the monitoring of the counts of the platelets represent the only valid predictive test of HELLP. These concerned women in the third trimester of pregnancy, especially those with a history of preeclampsia; 2. the treatment must be immediate, intensive and multidisciplinary, the plasmapheresis has remarkably improved the prognosis; 3. surgical treatment performed in order to control the SPPH makes use of packing, embolization and/or fastening of the common hepatic artery and, in extreme cases, total hepatectomy with transplantation. The Authors believe it is useful to suggest a national epidemiological research in order to estimate the real incidence of the syndrome in Italy and to establish the guidelines for the medico-surgical treatment.

Effects of ketoconazole on methylprednisolone pharmacokinetics and cortisol secretion.

The disposition of methylprednisolone was examined in six normal subjects after the injection of 20 mg iv methylprednisolone sodium succinate. Disposition studies were performed both without and with ketoconazole, 200 mg/day, for 6 days. Ketoconazole increased the methylprednisolone AUC and mean residence time (by 135% and 66%, respectively) and decreased clearance (60%), the terminal phase slope, and the volume of distribution. These findings are typical of macrolide antibiotic alteration of methylprednisolone disposition and consistent with reports of inhibition of drug metabolism by ketoconazole. Methylprednisolone reduced the 24-hour cortisol AUC by 44%, but morning cortisol concentrations returned to normal. Ketoconazole with methylprednisolone further reduced the 24-hour cortisol AUC and suppressed morning cortisol concentrations. Thus ketoconazole inhibits methylprednisolone disposition and extends the adrenal suppression effects of this corticosteroid.

Pharmacokinetics of single and multiple doses of ethinyl estradiol and levonorgestrel in relation to smoking.

The effects of tobacco and oral contraceptive (OC) use (Ovral) on the pharmacokinetics of levonorgestrel (0.25 mg) and ethinyl estradiol (50 micrograms) were examined. Young women (n = 27) were grouped as follows: I: non-OC users/nonsmokers; II: OC users/nonsmokers; III: non-OC users/smokers; and IV: OC users/smokers. The apparent clearance of levonorgestrel in group I was 80.9 +/- 15.6 ml/hr/kg and the half-life was 19.3 hours. A significant decrease in levonorgestrel clearance was seen in the chronic OC users (groups II and IV). The apparent oral clearance of ethinyl estradiol was 1002 +/- 398 ml/hr/kg in group I and the half-life averaged 7.7 hours. Groups II and III showed decreased (not significant) clearance of ethinyl estradiol. Tobacco use had no effect on steroid pharmacokinetics in the non-OC users. Although chronic OC use did not affect ethinyl estradiol clearance, a joint effect of tobacco/OC use on enhancing clearance of ethinyl estradiol appeared to occur. A linear relationship was found between 24-hour trough serum concentrations and AUC values of both steroids that may facilitate population monitoring studies of OC exposure.

PIP: The effects of combined cigarette smoking and oral contraceptive (OC) use on the pharmacokinetics of the pill's major components were examined in 27 white female volunteers grouped as follows: Group 1, non-OC user, nonsmoker; Group 2, OC user, nonsmoker; Group 3, non-OC user, smoker; Group 4, OC user, smoker. The 11 OC users in the study had been taking the pill for over 6 months; 5 were taking Ovral (50 mcg of ethinyl estradiol, 0.5 mg of norgestrel) and the remaining 6 switched to Ovral for the 1-month cycle before the study period. The non-OC users took 1 study dose of Ovral. The clearance of levonorgestrel was significantly lower in chronic OC users (mean elimination half-life of 30 hours) than in single-dose subjects (mean elimination half-life of 23 hours). The mean elimination half-life of ethinyl estradiol was approximately 12 hours for both chronic and acute OC use, although there was a nonsignificant tendency for lower ethinyl estradiol clearances in chronic OC users. Chronic tobacco use as a single factor did not influence the pharmacokinetics of levonorgestrel; however, a joint effect from chronic OC use and tobacco use was seen for ethinyl estradiol. Tobacco use had no effect on steroid pharmacokinetics in the non-OC users. Finally, a linear relationship was found between 24-hour trough serum concentrations and area-under-curve values of both steroids that may facilitate population monitoring studies of OC exposure.

Identification and isolation of human epithelial cell colonies that express specific gene products.

A new and simple method for the identification and isolation of human cell clones expressing specific gene products is presented. This technique is analogous to the colony blotting techniques described for prokaryotes. In this technique, human epithelial cells are grown in tissue-culture dishes, and cells from the colonies on the dish are transferred in situ to a high-density cationized quaternary amine-charged nylon membrane. The membrane can then be processed multiple times, using antibody and/or nucleic acid probes, for the identification of those cells producing the desired protein, mRNA or DNA sequence. Once identified, the desired cell colony is isolated for expansion, direct DNA sequencing and/or cell function. We demonstrate the potential of the technique by identifying and isolating colonies of replicating normal human liver hepatocytes producing albumin and keratin.

Prednisolone binding to albumin and transcortin in the presence of cortisol.

The protein binding of prednisolone was assessed in a 5% albumin solution and in pooled human serum, alone and in the presence of various amounts of cortisol. Significant displacement of prednisolone from transcortin binding sites occurred with little or no change in transcortin binding capacity or affinity constant for prednisolone, suggesting competitive inhibition of prednisolone binding by cortisol. Little or no displacement of prednisolone from the protein binding sites on albumin occurred though a decrease in the number of binding sites (four vs two), and an increase in the affinity constant for the albumin-prednisolone interaction (4.32 X 10(2) vs 9.43 X 10(2) M-1) occurred in the presence of cortisol. Allosteric conformational changes may occur in albumin structure in the presence of cortisol. These alterations have no effect on the fraction of prednisolone bound to albumin.

In vitro protein-binding characteristics of delavirdine and its N-dealkylated metabolite.

This study was performed to determine delavirdine protein-binding characteristics as well as those of its N-dealkylated metabolite (N-DLV). Initial studies of 36 microM delavirdine and 30 microM N-DLV in solutions of plasma, albumin 4 g%, alpha-1-acid glycoprotein (AAG) 100 mg% or immune globulin (IVIG) 5 g% were conducted. Delavirdine (12, 36 and 73 microM) and N-DLV (10, 30 and 60 microM) were then studied alone and in combination in plasma and various concentrations of albumin. Studies were done in triplicate using equilibrium dialysis. The mean delavirdine fraction unbound (fu) in plasma, albumin, IVIG and AAG was 0.013, 0.033, 0.752 and 0.912 while the mean fu of N-DLV in these same protein solutions was 0.139, 0.195, 0.329 and 0.359. In plasma and albumin, a greater fu was observed at higher delavirdine concentrations and no significant changes in fu were noted with the addition of N-DLV. An increase in delavirdine fu was noted as the albumin concentrations decreased. The fu of N-DLV increased significantly as the concentration of albumin decreased as well as with decreasing N-DLV concentration. The potential implications of extensive delavirdine binding to plasma proteins, primarily albumin, are discussed.

Drug compliance and unreported drugs in the elderly.

A random sample of 46 general practitioners of the Unitá Sanitaria Locale in Torino recruited 802 elderly outpatients and collected information about complaints and current drug treatment. Within a week each patient received a home interview and details were collected on drug compliance and use of drugs other than those reported by the general practitioners. On average, each patient was taking 3.6 drugs, of which 2.9 were correctly reported by the general practitioners and 0.7 were unreported. Among the most prescribed therapeutic groups there were drugs with a narrow therapeutic index (cardiovascular drugs, diuretics, psychotropic agents) and substances whose efficacy has never been fully documented ("cerebroactive-vasoactive" agents). Age and number of complaints were positively and significantly correlated with number of prescribed drugs. Nearly half of the sample (44.4%)--more frequently women and people with higher education--were taking one or more drugs not detected by the general practitioners, often benzodiazepines taken over a long period for anxiety or insomnia. Full compliance occurred for 81.5% of the prescriptions and 59.9% of patients were correctly taking all prescribed drugs. Compliance was lower for prescriptions of the general practitioners compared with other doctors' prescriptions (eg, hospital doctor, private doctor) and probability of taking correctly all the prescribed drugs decreased with the number of medicines concurrently taken. The most common reason for noncompliance was fear of side effects.

Increased vascular endothelial growth factor mRNA expression in the heart of streptozotocin-induced diabetic rats.

The aim of the present study was to evaluate vascular endothelial growth factor (VEGF), fms-like tyrosine kinase 1 (flt-1), and fetal liver kinase (flk-1) expression in the heart of experimental diabetic rats. Ten young adult male Wistar rats (5 streptozotocin [STZ]-induced diabetic rats, without insulin treatment, and 5 controls) were studied. Ninety days after the induction of diabetes, semiquantitative reverse transcription (RT)-polymerase chain reaction (PCR) coamplification of VEGF/glyceraldehyde 3-phosphate dehydrogenase (GAPDH) transcription was performed. RT-PCR was also performed for VEGF receptors flk-1 and flt-1. VEGF mRNA expression, at 234 bp, was detectable in the heart of the rats and was significantly higher in those with diabetes. Densitometric analysis of PCR products showed that VEGF mRNA levels were meanly 4.8-fold higher in STZ-induced diabetic rats than controls (VEGF/GAPDH densitometric ratio, 3.46 +/- 0.20 v 0.74 +/- 0.10, P <.001). No significant difference was found in flt-1 and flk-1 amplification products between STZ-induced diabetic rats and controls (flt-1/GAPDH densitometric ratio, 0.58 +/- 0.01 v 0.64 +/- 0.05, P>.1; flk-1/GAPDH densitometric ratio, 0.66 +/- 0.10 v 0.7 +/- 0.06, P >.2). The increase in VEGF mRNA expression observed in this experimental diabetic model is in contrast with the typical impairment in collateral vessels of diabetic hearts. This apparent discrepancy might be explained by a resistance of cardiac tissue to VEGF. The lack of mRNA flt-1 and flk-1 overexpression in diabetic hearts could be one of the mechanisms for this resistance.

Frequency-dependent changes in cerebral blood flow and evoked potentials during somatosensory stimulation in the rat.

Contrary to the concept of neuronal-vascular coupling, cortical evoked potentials do not always correlate with blood flow responses during somatosensory stimulation at changing stimulus rates. The goal of this study is to clarify the effects of stimulus frequency on the relationship between somatosensory evoked potentials (SEPs) and cerebral blood flow. In rats anesthetized with alpha-chloralose, we measured SEPs by signal-averaging field potentials recorded with an electrode placed on dura overlying the hindlimb somatosensory cortex. Regional blood flow was simultaneously assessed in the same region with a laser-Doppler flow (LDF) probe. The contralateral sciatic nerve was stimulated with 0.1 A pulses at the frequencies of 1, 2, 5, 10 and 20 Hz. SEPs (both P1 and N1 components) declined with increasing frequency regardless whether stimulus duration (20 s) or number (100) were kept constant, suggesting that frequency is an important determinant of neuronal activity. In contrast, LDF responses increased to a maximum at 5 Hz, and do not correlate with SEPs. Because CBF should reflect integrated neuronal activity, we computed the sum of SEPS (summation operatorSEP = SEP x stimulus frequency) as an index of total neuronal activity at each frequency. Summation operatorSEP indeed correlates positively (P<0.001) with LDF responses. Thus, during somatosensory stimulation at various frequencies, cerebral blood flow is coupled to integrated neuronal activity but not to averaged evoked potentials.

Selective loss of hippocampal long-term potentiation, but not depression, following fluid percussion injury.

We investigated the early effects of in vivo fluid percussion injury (FPI) on hippocampal synaptic potentials and excitability. In vitro field potential recordings and immunocytochemistry were performed in the CA1 region in slices from naïve, post-FPI, or sham-operated rats. The following electrophysiological and morphological parameters were affected following FPI: (1) threshold for population spike generation was increased suggesting that post-FPI neurons were hypoexcitable; (2) long-term potentiation (LTP) could not be induced in injured hippocampi; (3) GFAP and inducible NO synthase (iNOS) immunoreactivity were enhanced post-FPI; and (4) following injury, synaptophysin immunoreactivity was enhanced in CA1 stratum radiatum. The effects of FPI on synaptic plasticity were LTP-specific, since long-term depression (LTD) could be equally induced and maintained in post-FPI, sham-operated and control slices. Sham-operated slices were characterized by synaptic excitability indistinguishable from naïve controls, but displayed decreased ability for LTP production and expressed high levels of iNOS. We conclude that FPI causes a selective loss of LTP, possibly due to a previous potentiation induced by trauma as reflected by the increased expression of synaptic proteins. Sham surgical procedures were, however, not without effects on long-term potentiation itself; the latter effects appear to be mediated by an increased production of NO. Our study demonstrates for the first time that hippocampal slices can be used to investigate the correlates of in vivo FPI. Furthermore, we describe LTP-specific deficits in post-traumatic brain injury, suggesting that FPI can selectively erase one of the two main NMDA-dependent forms of synaptic plasticity in the hippocampus.

Mechanisms of nitric oxide-induced cytotoxicity in normal human hepatocytes.

Chronic exposure of hepatocytes to reactive nitrogen species (RNS) following liver injury and inflammation leads not only to functional and morphological alterations in the liver but also to degenerative liver diseases and hepatocellular carcinoma. Previously, we showed that S-nitroso-N-acetylpenicillamine-amine (SNAP), which generates nitric oxide, and 3-morpholinosydnonimine (Sin-1), which generates equal molar concentrations of superoxide and nitric oxide resulting in peroxynitrite production, exhibited different levels of cytotoxicity to normal human hepatocytes in culture. The aim of the present study was to elucidate some of the molecular and cellular pathways leading to hepatocyte cell death induced by RNS. Following treatment of the hepatocytes with SNAP or Sin-1, gene-specific DNA damage was measured in mtDNA and a hprt gene fragment using a quantitative Southern blot analysis. Both agents induced dose-dependent increases in DNA damage that was alkaline labile, but not sensitive to both formamidopyrimidine-DNA glycosylase (fpg) and endonuclease III, which recognize 8-oxoguanine, thymine glycol, and other oxidized pyrimidines. DNA damage was two- to fivefold greater in mtDNA than in the hprt gene fragment. There was a persistent and marked increase in DNA damage posttreatment that appeared to arise from the disruption of electron transport in the mitochondria, generating reactive species that saturated the repair system. DNA damage induced by Sin-1 and SNAP led to cell-cycle arrest in the S-phase, growth inhibition, and apoptosis. The data support the hypothesis that the functional and morphological changes observed in liver following chronic exposure to RNS are, in part, the result of persistent mitochondrial and nuclear DNA damage.

An enzyme-linked immunosorbent assay (ELISA) for the detection and quantitation of the tumor marker 1-methylinosine in human urine.

A highly sensitive enzyme-linked immunoassay (ELISA) was developed to detect and quantify the tumor marker, 1-methylinosine (m1I), in human urine. The rabbit antisera was highly specific for m1I with negligible or no inhibition by other nucleosides excreted into urine. Using the competitive ELISA, nanogram amounts of m1I were easily measured directly in urine. The assay agreed with our previous hplc analysis of m1I in urine for identifying those individuals with chronic myelogenous leukemia. Thus, this assay should greatly facilitate the quantitation of m1I as a tumor marker.

Differential response of normal, premalignant and malignant human oral epithelial cells to growth inhibition by chemopreventive agents.

Squamous cell carcinoma (SCC) of the oral cavity is a multistep process, progressing through a series of discrete, irreversible and complementary alterations in genes that control cell growth, death, and differentiation. In the premalignant state, the oral mucosa progresses through various grades of epithelial dysplasia, with the potential to convert to SCC. Chemopreventive strategies are designed to suppress, reverse, or prevent the formation of premalignant lesions and their subsequent progression to SCC. In the present study, we determined the growth inhibitory effect of 21 chemopreventive agents in a cell culture model using normal, premalignant, and malignant human oral mucosal cell lines. There were significant differences in the growth inhibitory responses of these cell lines to selected retinoids and non-retinoid analogs. Among the retinoids tested, the synthetic retinamides, as a class, showed selective growth inhibition of both premalignant and malignant cells compared to normal human oral epithelial cells in culture. Within the retinamide class, 2CPR exhibited the greatest selectivity in the growth inhibition of premalignant and malignant cells. Among the non-retinoids analyzed, DFMO was a moderate to potent inhibitor of malignant and premalignant oral cell growth, respectively, and stimulated normal oral cell growth at low concentrations. Using this in vitro approach, we have identified several potential chemopreventive agents for oral cancer as selective growth inhibitors of premalignant ahd malignant human oral mucosa cells.

Modulation of Ki67, p53 and RARbeta expression in normal, premalignant and malignant human oral epithelial cells by chemopreventive agents.

BACKGROUND: Aberrant expression of Ki67, p53 and RARbeta are characteristic of many tumor types including those of the oral cavity. Chemopreventive agents may act by modulating their expression to more normal levels. MATERIALS AND METHODS: The effects of 21 chemopreventive agents on the expression of Ki67, p53 and RARbeta were determined using a human in vitro model of normal, premalignant and malignant oral epithelial cell lines. RESULTS: Ki67 and mutant p53 (mtp53) were overexpressed in both the premalignant and malignant cell lines, whereas expression of RARbeta was high in the normal, low in the premalignant and not detectable in the malignant cell lines. Most of the agents selectively inhibited the expression of Ki67 in the premalignant and malignant cell lines. Eight of the 21 agents increased, while four agents decreased, the levels of mtp53 protein in the premalignant cell line. In the malignant cell line, five of the agents increased, while ten agents decreased mtp53 protein levels. The agents increased RARbeta expression to near normal levels in the premalignant cell line. CONCLUSION: The data suggest that the suppression of Ki67 and mtp53 are good indicators of the effectiveness of agents in premalignant and malignant oral cells, whereas the enhancement of RARbeta is a measure of effectiveness in premalignant oral cells.