Chronic Effects of Ethanol and/or Darunavir/Ritonavir on U937 Monocytic Cells: Regulation of Cytochrome P450 and Antioxidant Enzymes, Oxidative Stress, and Cytotoxicity.

BACKGROUND: Our recent study has shown that acute treatment with ethanol (EtOH) increases oxidative stress and cytotoxicity through cytochrome P450 2E1 (CYP2E1)-mediated pathway in U937 monocytic cells. U937 cells are derived from blood monocytes and are considered as the model system for HIV-related study. Since the prevalence of alcohol use in HIV-infected population is high, and HIV+ patients are on antiretroviral therapy (ART) soon after they are diagnosed, it is important to study the interactions between EtOH and ART in monocytes. METHODS: This study examined the chronic effects of EtOH and ART (darunavir/ritonavir), alone and in combination, on expression/levels of cytochrome P450 enzymes (CYPs), antioxidant enzymes (AOEs), reactive oxygen species (ROS), and cytotoxicity in U937 cells. The mRNA and protein levels were measured using quantitative reverse transcription polymerase chain reaction and Western blot, respectively. ROS and cytotoxicity were measured using flow cytometry and cell viability assay, respectively. RESULTS: While chronic ART treatment increased CYP2E1 protein expression by 2-fold, EtOH and EtOH+ART increased CYP2E1 by ~5-fold. In contrast, ART and EtOH treatments decreased CYP3A4 protein expression by 38 ± 17% and 74 ± 15%, respectively, and the combination additively decreased CYP3A4 level by 90 ± 8%. Expressions of superoxide dismutase 1 (SOD1) and peroxiredoxin (PRDX6) were decreased by both EtOH and ART, however, the expressions of SOD2 and catalase were unaltered. These results suggested increased EtOH metabolism, increased ART accumulation, and decreased defense against ROS. Therefore, we determined the effects of EtOH and ART on ROS and cytotoxicity. While ART showed a slight increase, EtOH and EtOH+ART displayed significant increase in ROS and cytotoxicity. Moreover, the combination showed additive effects on ROS and cytotoxicity. CONCLUSIONS: These results suggest that chronic EtOH, in the absence and presence of ART, increases ROS and cytotoxicity in monocytes, perhaps via CYP- and AOE-mediated pathways. This study has clinical implications in HIV+ alcohol users who are on ART.

Enhanced oxidative stress by alcohol use in HIV+ patients: possible involvement of cytochrome P450 2E1 and antioxidant enzymes.

BACKGROUND: Alcohol consumption is prevalent amongst HIV positive population. Importantly, chronic alcohol use is reported to exacerbate HIV pathogenesis. Although alcohol is known to increase oxidative stress, especially in the liver, there is no clinical evidence that alcohol increases oxidative stress in HIV positive patients. The mechanism by which alcohol increases oxidative stress in HIV positive patients is also unknown. METHODS: To examine the effects of alcohol use on oxidative stress we recruited HIV+ patients who reported mild-to-moderate alcohol use. Strict inclusion and exclusion criteria were applied to reduce the effect of other therapeutic drugs metabolized via the hepatic system as well as the effect of co-morbidities such as active tuberculosis on the interaction between alcohol and HIV infection, respectively. Blood samples were collected from HIV-negative alcohol-users and HIV positive alcohol-users followed by collection of plasma and isolation and fractionation of monocytes from peripheral blood. We then determined oxidative DNA damage, glutathione level, alcohol level, transcriptional level of cytochrome P450 2E1 (CYP2E1) and several antioxidant enzymes, and plasma level of cytokines. RESULTS: Compared to HIV-negative alcohol users, HIV-positive alcohol users demonstrated an increase in oxidative DNA damage in both plasma and CD14+ monocytes, as well as, a relative increase in oxidized/reduced glutathione (GSSG/GSH) in plasma samples. These results suggest an increase in oxidative stress in HIV-positive alcohol users compared with HIV-negative alcohol users. We also examined whether alcohol metabolism, perhaps by CYP2E1, and antioxidant enzymes are involved in alcohol-mediated increased oxidative stress in HIV-positive patients. The results showed a lower plasma alcohol level, which was associated with an increased level of CYP2E1 mRNA in monocytes, in HIV-positive alcohol users compared with HIV-negative alcohol users. Furthermore, the transcription of major antioxidants enzymes (catalase, SOD1, SOD2, GSTK1), and their transcription factor, Nrf2, were reduced in monocytes obtained from HIV positive alcohol users compared to the HIV-negative alcohol user group. However, no significant change in levels of five major cytokines/chemokines were observed between the two groups. CONCLUSIONS: The data suggests that alcohol increases oxidative stress in HIV+ patients, perhaps through CYP2E1- and antioxidant enzymes-mediated pathways. The enhanced oxidative stress is accompanied by a failure of cellular antioxidant mechanisms to maintain redox homeostasis. Overall, the enhanced oxidative stress in monocytes may exacerbate HIV pathogenesis in HIV positive alcohol users.

Evaluation of 1,2,5-thiadiazoles as modulators of M1/M5 muscarinic receptor subtypes.

Studies have demonstrated the presence of allosteric binding sites on each of the muscarinic acetylcholine receptor (mAChR) subtypes. Since most drugs targeting muscarinic receptors bind to the highly conserved orthosteric binding site, they fail to achieve appreciable subtype selectivity. Targeting non-conserved allosteric sites may provide a new way of enhancing selectivity for individual subtypes of muscarinic receptor. Tetra(ethyleneglycol)(3-methoxy-1,2,5-thiadiazol-4-yl)[3-(1-methyl-1,2,5,6-tetrahydropyrid-3-yl)-1,2,5-thiadiazol-4-yl] ether, CDD-0304 (10), was found to be a M1/2/4 selective muscarinic agonist and might prove useful in treating the symptoms associated with schizophrenia (J. Med. Chem.2003, 46, 4273). It was hypothesized that the observed subtype selectivity demonstrated by 10 may be due to its ability to function as a bitopic ligand (J. Med. Chem.2006, 49, 7518). To further investigate this possibility, a novel series of compounds was synthesized using a 1,2,5-thiadiazole moiety along with varying lengths of a polyethylene glycol linker and terminal groups, for evaluation as potential allosteric modulators of muscarinic receptors. Preliminary biological studies were performed using carbachol to stimulate M1 and M5 receptors. No significant agonist activity was observed at either M1 or M5 receptors for any of the compounds. Compound 18, 2-(4-methoxy-1,2,5-thiadiazol-3-yloxy)-N,N-dimethylethanamine fumarate (CDD-0361F) was found to block the effects of carbachol at M5 muscarinic receptors.

Effects of ceftriaxone on systemic and central expression of anti- and pro-inflammatory cytokines in alcohol-preferring (P) rats exposed to ethanol.

AIMS: Determine the effect of reduction in ethanol consumption by alcohol-preferring (P) rats, following ceftriaxone treatment, on the cytokines levels in prefrontal cortex (PFC) and plasma. METHODS: Following 5 weeks of free access to ethanol (15 and 30%), P rats were treated daily with ceftriaxone or saline vehicle for either 2 or 5 consecutive days. Plasma and PFC were collected from ceftriaxone- and saline vehicle-treated groups, and assayed for the levels of pro- and anti-inflammatory cytokines. RESULTS: A significant increase in the plasma level of anti-inflammatory cytokine IL-10 was observed in the ceftriaxone-treated group when compared with the saline-treated group in both the 2-day and 5-day treatments. Furthermore, ceftriaxone treatment for 2 days induced reduction in TNFα level in both plasma and PFC. Additionally, ceftriaxone treatment for 2 days significantly reduced the IFNγ level in PFC. CONCLUSION: These findings show the ability of ceftriaxone to reduce alcohol consumption and induce modulation of the anti-inflammatory and pro-inflammatory cytokines levels in P rats.

Leprosy and tuberculosis concomitant infection: a poorly understood, age-old relationship.

Historically, archaeological evidence, post-mortem findings and retrospective analysis of leprosy institutions' data demonstrates a high observed incidence of concomitant infection with leprosy and tuberculosis (TB). However, reports of concomitant infection in the modern literature remain scarce, with estimates of annual new case detection rates of concomitant infection at approximately 0.02 cases per 100,000 population. Whilst the mechanism for this apparent decline in concomitant infections remains unclear, further research on this topic has remained relatively neglected. Modelling of the interaction of the two organisms has suggested that the apparent decline in observations of concomitant infection may be due to the protective effects of cross immunity, whilst more recently others have questioned whether it is a more harmful relationship, predisposing towards increased host mortality. We review recent evidence, comparing it to previously held understanding on the epidemiological relationship and our own experience of concomitant infection. From this discussion, we highlight several under-investigated areas, which may lead to improvements in the future delivery of leprosy management and services, as well as enhance understanding in other fields of infection management. These include, a) highlighting the need for greater understanding of host immunogenetics involved in concomitant infection, b) whether prolonged courses of high dose steroids pre-dispose to TB infection? and, c) whether there is a risk of rifampicin resistance developing in leprosy patients treated in the face of undiagnosed TB and other infections? Longitudinal work is still required to characterise these temporal relationships further and add to the current paucity of literature on this subject matter.

Determinants of rural women's participation in India's National Leprosy Eradication Programme.

A multistage representative random sample of women and men from each of the 3 states of Bihar, Uttar Pradesh and West Bengal, from the rural blocks where the Leprosy Mission Hospitals were located were selected during 2010 to identify relevant factors that are preventing active participation of women and suggest corrective steps. Adult men and women were interviewed in depth, using a detailed checklist by the first author. A total of 1239 respondents 634 women and 605 men, were interviewed, only 44 women (7%) claimed that they had earlier participated in leprosy work, about 92% of the women felt that they had the potential to take part in leprosy work, and 70% showed willingness to participate. Factors that would encourage and facilitate more women to participate in leprosy work, included financial support (32.8%), convincing the family to grant permission (88%), and delegating them to work in proximity to their residences (15%). Some women respondents (11.0%) felt that they would provide their services voluntarily for social good. Women suggested that work should be delegated as per their capabilities and skills, and they should be given proper orientation, training and guidance. Hardly 5% of ASHA's in the clusters examined participated in leprosy related work, which needs stringent steps to re-orient and encourage them to undertake leprosy related work. It is concluded that rural Indian women are keen to play an important role in the national leprosy eradication program, with minimal support from the government and nongovernmental agencies in a truly community-based approach. This will benefit vast numbers of leprosy affected women as well as others.

Facial sensory loss in multi-bacillary leprosy patients.

Sensation over the face was estimated using the Semmes-Weinstein's monofilament (target force 0-05 gms) in a cohort of multi-bacillary (MB) patients whose clinical and ocular characteristics were available at the time of leprosy diagnosis. Among the 190 MB patients examined, 56 (30%) had areas of sensory impairment somewhere on the face and 43 (23%) had sensory deficit over the lids and/or the malar area. Lagophthalmos (adjusted OR 8.96, 95% CI 0.96-83.50), Type 1 reaction (aOR 2.47, 95% CI 1.11-5-52), history of reactions (aOR 6.36, 95% CI 2.40-16.85) and glove and stocking anaesthesia (aOR 3.49, 95% CI 1-40-8.70) were associated with impaired facial sensation. Hypoesthesia restricted to areas over the lids and/or malar area showed a stronger association with lagophthalmos (aOR 17.5, 95% CI 1.98-154.36). Loss of facial sensation appears to be associated with lagophthalmos in MB patients.

Peroneal strength as an indicator in selecting route of tibialis posterior transfer for foot drop correction in leprosy.

OBJECTIVES: Tibialis posterior tendon transfer (TPT) technique, using either the Circum-tibial (CT) or Inter-osseus (IO) route is the standard surgical technique to correct foot drop. The selection of the route of transfer is usually dependent on the surgeon's preference. This study aims to identify criteria to help make that selection. SUBJECTS AND METHODS: A study was carried out on 381 feet during the period 1999 to 2010. All the patients operated during this period were included in the study. The CT route was used for those with peronei power 4 or 5, while the IO route was used when peronei power was 3 or less. RESULTS: In this case series the mean effective range of motion (above 90 degrees) was 11 and 12 degrees in CT and IO routes, respectively. The results were comparable in terms of rest position, active dorsiflexion and effective range of motion. All patients had a post-operative heel to toe gait, except for one of the 381 operated feet. Only three of the 381 feet had a reduction in navicular height of more than 2 cm, the medial arch being maintained in the others. CONCLUSIONS: TPT is a standard procedure to correct foot drop deformity in leprosy. Pre-selection for route of transfer, CT or IO, based on peronei strength avoids the complication of iatrogenic inversion. The technique of insertion and routine tendo-achilles lengthening provides a good range of movement. The deep tunnelling has not compromised the results, while giving excellent cosmetic appearance.

A conceptual protocol for translational research in the complex reality of leprosy.

In response to an international strategy to prioritise and focus research efforts in leprosy, this conceptual protocol outlines a research plan to address key translational research priorities. The protocol describes in broad terms a five-phase psychosocial and service-related research programme to facilitate: prevention of delay in diagnosis, improvement of adherence with multi-drug therapy, the roll out of chemoprophylaxis and increased participation in community based rehabilitation. The protocol proposes a model of research utilisation and the notion of complexity to form an integrating theoretical framework for exploring and enhancing research translation. The proposed sequential research programme is characterised by traditional and participatory strategies, culminating in participatory implementation of findings. Publication of the conceptual protocol prior to operationalisation and commencement of the research aims to encourage debate, refinement of strategies, collaboration and the optimisation of resources.

Disability adjusted working life years (DAWLYs) of leprosy affected persons in India.

BACKGROUND & OBJECTIVES: Disability-adjusted life years (DALYs) have been accepted as a useful method to estimate the burden of disease, and can be adapted to determine the number of productive years lost due to the disability. DALY has been reported for many studies but not for leprosy. Hence this study was carried out in three States of India. In view of the fact that in this study, productive working years are used, the term is modified as DAWLY. METHODS: A representative random sample of 150 leprosy affected persons, 50 from each States of Uttar Pradesh, West Bengal and Chhattisgarh, was chosen, and data were collected on detailed work-life history, occupation, time when leprosy was discovered, reported and treatment started, break of job/loss of income due to leprosy. The loss of wages and durations were used to compute the life-years lost due to leprosy, and summarized over the average total duration of 42 years of productive work-life from 18 to 60 years. The percentage losses were determined and differences tested for statistical significance. RESULTS: The overall mean (± SE) disability adjusted working life years was 28.6 (±0.67), a reduction of 13.4 yr from the ideal productive working life period of 42 yr. The youngest patients with disability had a reduction of 41.4 per cent, as compared to the oldest patients. There was a significant increase in loss based on year for those whose disability started earlier (P=0.0024). INTERPRETATION & CONCLUSIONS: On an average, 30 per cent of the leprosy affected person's work life is lost due to disability.

Time trends in MB-PB ratio among untreated leprosy patients attending a referral hospital in UP, India during 2001 to 2010.

Secular trends in incidence of leprosy serve as a powerful tool in determining progress in reaching eradication. However, the interpretation of these trends must take into account both operational and epidemiological factors. A study was done to assess a time trend in the ratios of MB & PB from 2001 to 2010 based on the leprosy patients registered in a referral hospital in UP, India. Data were analyzed based on the gender, age and residence. Regardless of these factors, MB proportion shows no significant trends. These findings are discussed and it is concluded that we are no more close to eradication as compared to the status over a decade ago. Hence, much greater efforts will be required to promote early detections of MB cases, whether children or adults, male or female.

Nursing interventions to manage anxiety levels of female inpatients admitted first time in a leprosy hospital.

Leprosy causes not just physical disabilities but mental and psycho social problems which are further more enhanced in women due to their submissive and secondary role in an Indian culture. This is reflected in their reluctance and delay in seeking hospitalization and generates great anxiety while admitted as inpatients. Appropriate nursing care can relieve much anxiety and help in faster healing. This paper presents the findings from such research carried out at a leprosy referral hospitalin north India. Adult female leprosy patients newly admitted for the first time in a leprosy referral hospital were interviewed in depth using Hamilton Anxiety Rating Scale and observed before and after implementing a customized nursing care plan. On admission, out of 40 women admitted more than 80% showed moderate or severe anxiety. After well planned nursing interventions only 2 continued to have severe anxiety, and a majority in all age groups showed significant reductions in anxiety levels, and responded well to leprosy care at the hospital. Well planned nursing care reduces or minimizes anxiety levels of female leprosy patients admitted first time in the hospital, and should become a standard practice in all hospital admissions.

Impact of COVID-19 pandemic on cross-country runners: changes in training, sleep, diet, and mental health.

Objective: Determine the impact of the COVID-19 pandemic on the changes in training, sleep, diet, and mental health of cross-country runners. Participants: Cross-country runners from NCAA Division II institutes. Methods: Wilcoxon signed-rank test was performed to analyze survey responses to ordinal questions on the survey while Spearman's rank correlation analysis (ρ) was used to calculate correlation between after the start of pandemic questions. Data was marked significant at p < 0.05. Results: Analysis of the survey responses revealed that cross-country runners were more likely to experience feelings of depression (p < 0.001), lack of motivation (p < 0.001), and higher daily stress (p < 0.001) after the start of the pandemic. After the start of the pandemic, runners running less days per week were more likely to report an increased feeling of depression (ρ=-0.315, p = 0.008) and lack of motivation (ρ=-0.458, p < 0.001). Conclusions: The present study underscores the importance of training, sleep, diet, and mental health amongst cross-country runners.

Help-seeking habits of untreated leprosy patients reporting to a referral hospital in Uttar Pradesh, India.

In a potentially disabling disease like leprosy, the need for prompt diagnosis and start of treatment cannot be over-emphasized. With the advent of massive Information, Education and Communication (IEC) strategies and easy accessibility of free multi drug therapy (MDT), delay in treatment is more dependent on patient initiative and subsequent health seeking habits. To study the factors contributing to delay, a random sample of 86 new untreated leprosy patients presenting to The Leprosy Mission Community Hospital, Naini, Allahabad during 2011 were interviewed in depth with the help of a check list. 61% of patients had disability at first presentation. The most common first symptom was a hypopigmented patch. Mean delay was found to be 25.9 months. Reasons for delay varied from ignorance aboutthe symptoms and signs of the disease, monitoring of symptoms in the hope that they would disappear by themselves and lack of vigilance among local medical practitioners in the lower levels of the health system. The authors discuss the typical sequence of events that contributed to delay at each stage before finally presenting at a referral hospital. It is necessary to outline recommendations to address delay in terms of intense health education campaigns, mass communication strategies and developing a high index of suspicion among primary health care givers.

Nutritional status of leprosy patients in India.

Across-sectional epidemiological study was carried out at a Leprosy Referral Hospital in Delhi to assess the nutritional status of multibacillary leprosy patients in comparison to the general population using BMI. 150 people affected with multibacillary leprosy were included in the study, of whom 108 (72%) had WHO Grade 2 disability. 100 non leprosy patients were also included as a control group. Socio-demographic and clinical details as well as their height and weight were measured and the BMI computed. The findings clearly showed that under-nutrition (BMI < 18.5) was more common in people affected by leprosy than in those without leprosy, regardless of age or sex. Presence of disability made the incidence of under-nutrition more likely. The duration of disease, number of lesions or bacterial index had no impact on the level of nutrition. There may be multiple factors working together to lead to this under-nutrition and these are discussed briefly. If, we aim to provide high quality services with a holistic approach, a mandatory BMI should be calculated for every patient and if under nourished, a qualitative diet summary should be done and suitable nutritional advice given. Further, studies are needed for a better understanding of the occurrence and progression of under-nutrition in leprosy to find efficient ways to combat this problem.

Efficacy of temporalis muscle transfer for correction of lagophthalmos in leprosy.

OBJECTIVES: Temporalis Muscle Transfer (TMT) is a surgical technique used to correct lagophthalmos in leprosy patients. We have evaluated the degree of success of TMT in achieving full lid closure, which is important in preventing damage to the cornea. SUBJECTS AND METHODS: A retrospective study was carried out on 69 patients who had TMT done, at one centre, on 101 eyes during the period of 1998-2009. Lid gaps on direct gaze and with both gentle and forced closure, as well as voluntary muscle testing of eye lid closure, were assessed using standard measuring techniques by a qualified physiotherapist. Associated problems due to lagophthalmos were recorded both pre- and post- operatively. Data were abstracted on to a special proforma and subjected to statistical analysis using SPSS. RESULTS: On completion of post-operative physiotherapy, 85% of the eyes could achieve full lid closure with no measurable gap. The mean (SD) lid gap on forced closure was 48 (2.8) mm pre-operatively and 0.2 (0.5) mm at the end of the in-patient stay. The mean (SD) lid gap on gentle closure was 7.9 (2.6) mm preoperatively and 2.4 (1.8) mm post-operatively. The mean (SD) vertical inter-palpebral distance, during straight gaze, was reduced from 12.6 (1.6) pre-operatively to 9.8 (1.2) postoperatively. Exposure keratitis cleared in 16 of 27 eyes (60%) and Epiphora cleared or improved in 31 eyes. CONCLUSIONS: It is concluded that the TMT is a successful option (cosmetically and functionally) for correction of lagophthalmos.

Sustainability of integrated leprosy services in rural India: perceptions of community leaders in Uttar Pradesh.

As part of a community-based action research to reduce leprosy stigma, village committees were formed in 3 hyper endemic states of India. From a total of 10 village committees with nearly 200 members from Uttar Pradesh, a systematic random sample of 69 men and 23 women were interviewed in-depth regarding their views on sustainability of integrated leprosy services, as currently adopted. Their recommendations were also sought for further enhancement. Percentages were computed and compared for statistical significance using the z-normal test. The findings show that less than 50% of the respondents were confident that the present trend in voluntary early reporting for MDT and management of complications was adequate to sustain the integrated leprosy services. There were no differences by men or women members and they felt that lack of proper facilities, training and orientation of staff are most influencing factors. Many suggestions were given for improving the sustainability.

Medical and social concerns of leprosy cured after integration in India.

In order to understand nature of the medical and society related problems of leprosy cured individuals from various socio-cultural groups and develop proper system to address the current needs of leprosy cured, data have been collected from leprosy cured, employing qualitative methods such as FGDs, open ended interviews and participant observation and a quantitative survey method. The findings show that there has been not much change in the socio-cultural and economic situation of the leprosy cured especially the disabled ones, except a recognition in the health system that their anti-leprosy treatment is completed. In the present integrated health care system, disabled leprosy patients are also expected to seek their own health care by themselves along with general public, which is not happening because the leprosy cured are not yet perceived to be cured of leprosy by community as well as by themselves. Though some of the problems of leprosy cured seem to be poverty related, qualitative analysis shows, a vast gap between poverty problem of leprosy afflicted and non leprosy afflicted. Any special programme to bridge the gap between PHC and leprosy community is required and the new functionaries like ASHA in National Rural Health Mission (NRHM) and similar developments could be of vital use to make them cured in totality.

Leprosy among tribal population of Chhattisgarh state, India.

Chhattisgarh state is still endemic for leprosy and has a large tribal population. During 2003-2009, a total of 1530 untreated leprosy cases reported to the Leprosy Mission Referral Hospital in Champa, of which 151(9%) were classified as belonging to the scheduled tribes. The characteristics of these new tribal patients are described and compared with other patients and to the demographics of the tribals in the general population of the State. While tribals were accessing the leprosy services similar to the other social groups, the delay in reporting, high BI and other features pose more serious problems in the transmission of leprosy and in management of complications due to the harsh environment and occupational patterns of the tribals. Appropriate strategies and more community based approaches will be necessary if these groups are also targeted for eradication of leprosy.

Public awareness on integration of leprosy services at primary health centres in Uttar Pradesh, India.

Leprosy services were integrated into the general health a decade ago but it seems that a majority of public are still ignorant of this development. Hence, a study was done in Uttar Pradesh, India to determine the awareness about integration and its relationships to various socio-demographic factors. A multistage representative random sample of 3000 persons was chosen in Faizabad district, selecting a sample of 3 villages each situated within 1 km, 1-3 km and beyond 3 km of a PHC. A systematic random sample of 10% of households was chosen from selected villages and an adult male and an adult female from each household interviewed by a qualified investigator. Data were computerized and cross- tabulated against distance from the PHC, sex, age, education and occupational status. Only 45.7% in Uttar Pradesh are aware of the availability of leprosy treatment facilities at PHC but most knew that MDT was free. A smaller proportion was also aware of other facilities such as ulcer dressing and treatment of complications. Family members and health workers and PHC were the main source of information. It is concluded that massive efforts are urgently needed to educate the rural public on integration.