A rat model of polymicrobial infection in full-thickness excision wounds.

AIM: A reproducible animal model is required to study the pathophysiology of wound infections and for development of effective therapeutic interventions. The objective of this study was to produce an infected skin wound model utilizing the cecal microbiota in non-immunocompromised rats. MATERIALS AND METHODS: An excision wound was created on the dorsal surface of rats and inoculated with different concentration of cecal slurry (CS). Wound progression was investigated macroscopically by wound scoring and imaging. The rats were sacrificed on day 6 and microbial load, myeloperoxidase activity, histopathology, and scanning electron microscopy (SEM) were performed in wound tissue. RESULTS: Inoculation of CS into excision wounds caused significantly (p < 0.05) delayed wound healing in comparison to non-infected wounds as revealed by slow wound closure (9.1 to 12.83%). A significant (p < 0.05) difference in wound score was observed between the infected and non-infected wounds. A significantly (p < 0.05) high microbial load (~10 9 CFU/gm) was observed in infected wound which was supported by the presence of intensive bacterial colonization with sparse development of amorphous material on wound tissue during SEM analysis. A maximum increase of 1.76-fold in myeloperoxidase activity was observed in the infected wounds in comparison to non-infected wounds. Histopathology revealed increased amount of cellular infiltration, hematoma formation, and presence of bacterial aggregates in deep tissues. CONCLUSION: The study reports a reproducible and relevant clinical model of wound infection where cecal microbiota was used as a source of infection. This model can provide a suitable platform for evaluation of new therapeutic interventions.

Prevalence and predictors of self-reported risk behaviors among male injecting drug users.

BACKGROUND: Unsafe injecting drug use is a documented risk factor for the transmission of human immunodeficiency virus infection. Harm reduction strategy aims at reducing this deleterious consequence. OBJECTIVES: To study the prevalence and predictors of injecting as well as sexual risk behavior among male injecting drug users (IDUs). METHODS: A cross-sectional study was conducted among 250 male IDUs from January 2017 to March 2018. Risk behavioral data were collected after obtaining informed consent of the study participants. This included information on sharing, reusing needles/injections, and sexual behavior. Descriptive statistics and logistic regression model was run using the Epi Info software version 7.2 for Windows. RESULTS: In the past one month, 25.3% had shared needles/syringes by either borrowing or lending or both. Inconsistent condom use during sexual intercourse with a regular, casual, and paid sex partner was practiced by 64.7% (77/119), 65.8% (48/73), and 45.0% (18/40), respectively. The binary logistic regression analysis revealed that unsafe injecting drug behavior was higher among daily users: (OR = 3.0 [1.3-6.6]) and comparatively lower among those who preferred to avail needles/syringes from Needle Syringe Exchange Program (OR = 0.4 [0.2-0.9]); as compared to their counterparts. CONCLUSIONS: The findings suggest that IDUs in the study area are engaging in risk behaviors. Behavior change communication and harm reduction strategy should be strengthened.

A standardized Hippophae extract (SBL-1) counters neuronal tissue injuries and changes in neurotransmitters: implications in radiation protection.

CONTEXT: Effects of a radioprotective, standardized leaf extract (code SBL-1) from traditional medicinal plant, sea buckthorn [Hippophae rhamnoides L. (Elaeagnaceae)], on neurotransmitters and brain injuries in rats showing radiation-induced conditioned taste aversion (CTA), are not known. Understanding CTA in rats is important because its process is considered parallel to nausea and vomiting in humans. OBJECTIVE: This study investigated the levels of neurotransmitters, antioxidant defences and histological changes in rats showing radiation CTA, and their modification by SBL-1. MATERIALS AND METHODS: The inbred male Sprague-Dawley rats (age 65 days, weighing 190 ± 10 g) were used. Saccharin-preferring rats were selected using standard procedure and divided into groups. Group I (untreated control) was administered sterile water, group II was 60Co-γ-irradiated (2 Gy), and group III was administered SBL-1 before irradiation. Observations were recorded up to day 5. RESULTS: Irradiation (2 Gy) caused (i) non-recoverable CTA (≥ 64.7 ± 5.0%); (ii) degenerative changes in cerebral cortex, amygdala and hippocampus; (iii) increases in brain dopamine (DA, 63.4%), norepinephrine (NE, 157%), epinephrine (E, 233%), plasma NE (103%) and E (160%); and (iv) decreases in brain superoxide dismutase (67%), catalase (60%) and glutathione (51%). SBL-1 treatment (12 mg/kg body weight) 30 min before irradiation (i) countered brain injuries, (ii) reduced CTA (38.7 ± 3.0%, day 1) and (iii) normalized brain DA, NE, E, superoxide dismutase, catalase and CTA from day 3 onwards. DISCUSSION AND CONCLUSION: Radiation CTA was coupled with brain injuries, disturbances in neurotransmitters and antioxidant defences. SBL-1 pretreatment countered these disturbances, indicating neuroprotective action.

Ninety days of repeated gavage administration of Rhodiola imbricata extract in rats.

Rhodiola imbricata is a high-altitude plant, possesses adaptogenic, immunomodulatory, anti-oxidant and cytoprotective activity, and is widely used in traditional medicine. The present study was designed to ascertain the safety of aqueous extract of R. imbricata root when administered by gavage to rats for 90 days. Four groups of animals, each consisting of 15 males and 15 females, were administered 0, 100, 250 or 500 mg kg(-1) extract, in a single dose per day. The experimental rats when administered 100 mg kg(-1) of extract did not show any significant change in their body weight gain, organ/body weight ratio, or histological, hematological and biochemical variables studied. However, at higher doses of 250 and 500 mg kg(-1) extract, an increase in the body weight of rats of both the sexes was apparent without any change in their organ/body weight ratio. Furthermore, a noteworthy increase in plasma glucose and protein levels was recorded at both the higher doses, which were restored to normal after a 2-week withdrawal of treatment. Based on the findings of this study, the no observed effect level was 100 mg kg(-1) body weight per day of aqueous root extract of R. imbricata in rats administered subchronically.

Mental health, economic well-being and health care access amid the COVID-19 pandemic: a mixed methods study among urban men who have sex with men in India.

Scant empirical research from Asia has addressed the impact of COVID-19 on sexual minority health. We aimed to explore and understand the impact of COVID-19 on income security, mental health, HIV risk and access to health services among men who have sex with men (MSM) in India. We conducted a concurrent mixed methods study from April to June 2020, including a cross-sectional survey and in-depth semi-structured interviews with MSM recruited from three non-governmental organisations providing HIV prevention services in Chandigarh, India. We examined the associations of sexual minority stressors (sexual stigma, internalised homonegativity), economic stressors, and stress due to social distancing, with depression and anxiety, HIV risk, and access to health services. Survey findings (n = 132) indicated that internalised homonegativity and stress related to social distancing were significantly associated with depressive and anxiety symptoms. Results also showed reduced access to condoms, HIV testing and counselling services. Qualitative findings (n = 10) highlighted adverse economic impacts of COVID-19, including loss of employment/wages and engaging in survival sex work, which contributed to psychological distress and HIV risk. The COVID-19 pandemic has resulted in considerable psychological and financial distress among low socioeconomic status MSM in India, including those involved in sex work - communities already marginalised in economic, family and healthcare sectors. Structural interventions to improve access to mental health and HIV services and decrease financial burden are critical to mitigate the impact of COVID-19.

A retrospective analysis of sexually transmitted infections among males presenting to a tertiary care hospital of India.

Context: Sexually transmitted infections (STIs) are one of the most neglected diseases, leading to a high percentage of morbidity and mortality in India. The World Health Organization estimated that 20% of persons living with human immunodeficiency virus/acquired immunodeficiency syndrome are in their 20s and one out of twenty adolescents contract an STI each year. Aims: The present study was conducted to study the characteristics of the pattern of STI in adult males and study the prevalence of various STIs among them. Settings and Design: This retrospective study was conducted by retrieving records of males presenting to STI laboratory of our tertiary care hospital between (April 2018 and December 2019). Subjects and Methods: The patients comprised high-risk group males, approached through nongovernmental organizations (NGOs) and slum population visiting the dispensary attached to our institute. The age group of the patient included was between 0 and 85 years. Results: A total of 1023 males presented to our STI laboratory out of which 124 (12.12%) were symptomatic. The most common complaint was urethral irritation seen in 22.5%, followed by discharge in 9.6%. The most common sexually transmitted disease among symptomatic (34/124) as well as asymptomatic (172/899) men was syphilis showing a combined prevalence of 20% (206/1023). Out of 124 symptomatic patients, 29 (23.38%) complained of urethritis due to gonococcal infection. The association between the two was found to be significant (i.e., P < 0.05). Conclusion: STIs are a serious health problem in our country. Approximately 6% of the adult population have one or the other STI amounting to 30-35 million cases per year. An intensive study is the need of the hour which could help clinicians as well as microbiologists to control the spread of these infections.

Prevalence of nonviral reproductive tract infections/sexually transmitted infections in female patients with cervicovaginal discharge: Excerpts from a regional reference center in North India.

Background: To study the prevalence of common nonviral reproductive tract infections/sexually transmitted infections (RTI/STI) prevailing among females who presented to our regional STI reference center and to ascertain the association of various symptoms with different RTI/STIs. Materials and Methods: A retrospective analysis of female patients presenting to our STI Regional center located in the Department of Medical Microbiology in PGIMER, Chandigarh, was done between April 2018 and December 2019 for patients presenting with cervico-vaginal discharge. Two to three swabs were collected from each patient. The first swab was subjected to wet mount, gram stain, Potassium hydroxide (KOH) test, and culture on blood agar, the colonies obtained were identified by matrix-assisted laser desorption time of flight mass spectrometer (MALDI TOF-MS). Second swab was used for DNA extraction and detection of Neisseria gonorrhoeae (NG), Trichomonas vaginalis (TV), and Chlamydia trachomatis (CT) by polymerase chain reaction. The third swab, when available, was inoculated onto pleuropneumonia-like organisms (PPLO) broth. Results: One thousand and thirteenth of 1472 (69%) female patients were symptomatic and the most common presenting symptoms were vaginal discharge (707/1013 [69.8%]), infertility (266/1013 [26.2%]), genital itching (60/1013 [5.9%]), lower abdomen pain (47/1013 [4.6%]) and burning micturition (16/1013 [1.6%]). The most prevalent RTI/STI was bacterial vaginosis (BV) 18.2% (269/1472), followed by vulvovaginal candidiasis (VVC) 6.8% (100/1472) and trichomoniasis (TV) 1.9% (28/1472). Five cases each of Mycoplasma genitalium and Ureaplasma urealyticum, three of NG and one of CT were also identified. Coinfections were seen in 40 (2.7%) cases. The most common causative agent responsible for VVC in our study was Candida albicans (65%). Conclusion: RTI/STIs were common among women and 69% were symptomatic. BV was the most common STI present in 18.2%, followed by VVC (6.8%) and trichomoniasis (1.9%).

Yoga as a Preventive Intervention for Cardiovascular Diseases and Associated Comorbidities: Open-Label Single Arm Study.

Aim: Common Yoga Protocol (CYP) is a standardized yoga protocol authored by experts from all over the world under the aegis of the Ministry of AYUSH, Ayurveda, Yoga and Naturopathy, Unani, Siddha, Sowa Rigpa and Homeopathy (AYUSH). The potential of CYP can be determined as a cost-effective lifestyle modification to prevent the risk of developing cardiovascular diseases (CVD). Methods: In this prospective trial, we compared the effect of CYP at baseline and after 1 month. A total of 374 yoga-naïve participants performed CYP under the supervision of experienced trainers. Physiological [body mass index (BMI), blood pressure, percent oxygen saturation], biochemical (fasting blood glucose and lipid profile), and neurocognitive parameters were measured before and after the intervention. Results: At day 30 of yoga practice, serum levels of low-density lipoprotein (LDL), total cholesterol (TC), and high-density lipoprotein (HDL) were found significantly improved as compared to the baseline levels observed at the time of enrollment. Similarly, the lipid profile was also obtained from experienced trainers and found to be significantly different from those of yoga-naïve volunteers. When the intervention was compared between the healthy yoga-naïve participants with yoga-naïve participants suffering from medical issues, it was found that cholesterol profile improved significantly in the healthy-naive group as compared to the diseased group (hypertension, diabetes, underwent surgery, and CVD). Conclusion: These results highlight the need for further research to better understand the effects of yoga on the primary prevention of CVD.

Podophyllum hexandrum-Mediated Survival Protection and Restoration of Other Cellular Injuries in Lethally Irradiated Mice.

This study aims at the development of a safe and effective formulation to counter the effects of lethal irradiation. The sub-fraction (G-001M), prepared from Podophyllum hexandrum has rendered high degree of survival (>90%) at a dose of 6 mg kg(-1) body weight (intramuscular) in lethally irradiated mice. Therapeutic dose of G-001M, at about 20 times lower concentration than its LD(100), has revealed a DRF of 1.62. Comet assay studies in peripheral blood leukocytes have reflected that, treatment of G-001M before irradiation has significantly reduced DNA tail length (P < .001) and DNA damage score (P < .001), as compared to radiation-only group. Spleen cell counts in irradiated animals had declined drastically at the very first day of exposure, and the fall continued till the 5th day (P < .001). In the treated irradiated groups, there was a steep reduction in the counts initially, but this phase did not prolong. More than 60% decline in thymocytes of irradiated group animals was registered at 5 h of irradiation when compared with controls, and the fall progressed further downwards with the similar pace till 5th day of exposure (P < .001). At later intervals, thymus was found fully regressed. In G-001M pre-treated irradiated groups also, thymocytes decreased till the 5th day but thereafter rejuvenated and within 30 days of treatment the values were close to normal. Current studies have explicitly indicated that, G-001M in very small doses has not only rendered high survivability in lethally irradiated mice, but also protected their cellular DNA, besides supporting fast replenishment of the immune system.

Topical application of Jatyadi Ghrita and Jatyadi Taila accelerates wound healing in Sprague-Dawley rats: a study in gamma-radiation-induced skin wound model.

PURPOSE: Radiation-induced skin wounds/dermatitis can occur due to therapeutic, occupational, or accidental exposure to ionizing radiation. This study investigated the therapeutic efficacy of standardized Ayurvedic formulations [Jatyadi ghrita (JG) and Jatyadi taila (JT)] against 60Co-γ-radiation-induced acute skin wounds in rats. MATERIAL AND METHODS: Animal's [Sprague-Dawley rats (200 ± 20 g)] flanked skin was locally exposed to 45 Gy radiation (R45Gy) in Cobalt-60-teletherapy unit (Bhabhatron) to generate radiation wounds. JG and JT were applied topically twice daily on wounds from day 14 onwards after appearance of moist desquamation and wound healing efficacy was observed for a period of 42 days. RESULTS: R45Gy induced significant time dependent changes in rat's skin with erythema on day 7 followed by dry and moist desquamation. JG and JT application significantly (p < .001) reduced skin damage score, wound area (92% and 97% respectively on day 42), and bacterial load, when compared with R45Gy and showed better efficacy than sucralfate and betamethasone (positive controls). Formulations significantly reduced lipid peroxidation and enhanced antioxidant defenses, reduced inflammatory infiltrates and collagen fibers deposition as evident by decreased myeloperoxidase and hydroxyproline levels, and also reduced transforming growth factor-beta 1 (TGF-ß1) expression. Further, histology revealed reduced epidermal hyperplasia and dermal thinning with improved densities of hair follicles. Formulations were found to be nontoxic on 28 days application. CONCLUSIONS: The results demonstrated that JG and JT accelerated wound healing in irradiated skin tissue by faster re-epithelialization; reducing inflammation, collagen fibers deposition, and TGF-ß1 expression, indicated their potential human application in countering radiation wounds.

Whole genome exon arrays identify differential expression of alternatively spliced, cancer-related genes in lung cancer.

Alternative processing of pre-mRNA transcripts is a major source of protein diversity in eukaryotes and has been implicated in several disease processes including cancer. In this study we have performed a genome wide analysis of alternative splicing events in lung adenocarcinoma. We found that 2369 of the 17 800 core Refseq genes appear to have alternative transcripts that are differentially expressed in lung adenocarcinoma versus normal. According to their known functions the largest subset of these genes (30.8%) is believed to be cancer related. Detailed analysis was performed for several genes using PCR, quantitative RT-PCR and DNA sequencing. We found overexpression of ERG variant 2 but not variant 1 in lung tumors and overexpression of CEACAM1 variant 1 but not variant 2 in lung tumors but not in breast or colon tumors. We also identified a novel, overexpressed variant of CDH3 and verified the existence and overexpression of a novel variant of P16 transcribed from the CDKN2A locus. These findings demonstrate how analysis of alternative pre-mRNA processing can shed additional light on differences between tumors and normal tissues as well as between different tumor types. Such studies may lead to the development of additional tools for tumor diagnosis, prognosis and therapy.

Questionnaire based counselling: A tool to identify stress precipitating factors and thereby upgrading the performance of a new medical Entrant in department of anatomy.

BACKGROUND AND AIMS: Vast first MBBS course has been posing great problems for a fresh entrant in medical college. A student is merely 18-19 years when he gains entry to a medical college in India. Multiple problems pile up for him at the very onset of his academic year in Department of Anatomy. The present study aimed to identify various stress factors and further assess the effect of counselling/mentorship programme on solving such issues. MATERIALS AND METHODS: A pre-validated, predesigned questionnaire was administered to hundred first year medical students. They were individually counselled with various faculty persons. Based on the questionnaire given, the results were analysed. RESULTS: 45% of the students were sure that their language problem was solved after the session. 48% students accepted that they did have some adjustment problems, which were solved then. 30% expressed that some of their personal problems were also solved. 54% were now able to manage their time judiciously. 64% opined that counselling helped to plan their studies well. 52% students admitted that counselling improved their performance in theory. 46% expressed that counselling also helped to improve their performance in practicals. 65% told us that their skills both in theory and practical had improved after the session. 39% consented that counselling relieved them of anxiety. 67% recognised that their knowledge in the subject of anatomy had increased. 75% were of the view that similar sessions should be conducted more frequently. 40% preferred to express their views in their regional language. 85% students advised us to carry forward these sessions to the next batches. CONCLUSION: Our study highlights the need for counselling to solve multifaceted adjustment problems.

Assessment of newly introduced foundation course for medical undergraduates: students' vs faculty's perspective.

BACKGROUND AND OBJECTIVES: In compliance with MCI's recommendation, a month-long Foundation Course was conducted in our institution in Aug'19. The present study was conducted to seek the opinion of the students and faculty regarding relevance of the topics included in the course to enable revisions to be made in designing the course for the subsequent sessions, to make it more effective and student-oriented. METHODS: A questionnaire-based cross-sectional study was conducted to seek the opinion of 100 students who underwent the Foundation Course and 35 faculty members who were involved in teaching the topics allotted. RESULTS: Positive feedback was recorded from63% of the students and 69% of the faculty regarding the overall experience of the course. Among the different modules, Skills module was voted as relevant by majority (73%) of students, whereas a module on Enhancement of Language and Computer Skills was found relevant to the course by a mere 52% students. In contrast, module on Sports and Extracurricular activities was adjudged as most relevant while as module on Enhancement of Language and Computer Skills got the least positive feedback (81.43% vs. 60.36%, respectively) from the faculty. INTERPRETATION AND CONCLUSIONS: Foundation Course recommended by MCI for MBBS students at the entry level is a welcome step. This can further be made more beneficial by making necessary modifications in the planning of the course in light of suggestions received from the participants.

Search for a functional glucocorticoid receptor in the mammalian lens.

Prolonged glucocorticoid treatment of medical conditions such as rheumatoid arthritis or asthma can lead to the formation of a posterior subcapsular cataract as a negative side effect. Currently, the only treatment for this cataract is surgery because very little is known about the mechanism of glucocorticoid action in the mammalian lens. Understanding of a lens glucocorticoid response is essential for the treatment and prevention of a steroid induced cataract. It has been suggested that glucocorticoids exert their effects on the lens indirectly, non-specifically, or through non-classical mechanisms. While these modes of action may contribute to the formation of glucocorticoid induced posterior subcapsular cataract, the finding of a classical, specific, functional lens glucocorticoid receptor suggests that glucocorticoids target lens epithelial cells directly, specifically, and similar to what has been observed in other cells types. This review explores the discovery of the glucocorticoid receptor in humans lens epithelial cells and the lens specific glucocorticoid response. The distinct changes in lens epithelial cell signaling pathways (MAPK and PI3K-AKT) suggest that glucocorticoids modulate several cellular functions and may explain why a lens glucocorticoid response has been difficult to elucidate.

Association between CCND1 G/A870 polymorphism, allele-specific amplification, cyclin D1 expression, and survival in esophageal and lung carcinoma.

PURPOSE: Cyclin D1 is found on 11q13, which is a region frequently amplified in several tumor types. The CCND1 locus gives rise to at least two protein isoforms of D1 (D1a and D1b). A common G/A polymorphism (G/A870) is thought to influence the expression levels of D1a and D1b. D1b has been suggested to be increased in the presence of the A allele and more oncogenic than D1a. Furthermore, the A allele has been reported to correlate with increased risk of carcinoma in several tumor types, suggesting that this polymorphism and D1b are important in tumor progression. However, contradictory data about the polymorphism, D1 variant expression, and correlation with survival have been reported. We explored the relationship between gene amplification, G/A870 genotype, D1a and D1b expression, and overall survival in esophageal adenocarcinoma and non-small cell lung cancer. EXPERIMENTAL DESIGN: DNA and RNA were isolated from 54 esophageal adenocarcinoma samples and 89 non-small cell lung cancer samples and were analyzed for gene amplification, genotype at the polymorphism, gene expression, and association with overall survival. RESULTS: The D1 variant expression did not correlate with amplification, genotype, or overall survival in either tumor type. The total D1 expression correlated with decreased patient survival. Several other genes on 11q13 also seem to be overexpressed and correlated with decreased survival. CONCLUSIONS: We report that the G/A870 polymorphism does not correlate with patient survival, or with D1a or D1b expression. However, the total D1 expression and the expression of several other genes on 11q13 seem to be associated with esophageal adenocarcinoma patient survival.

Specific activation of the glucocorticoid receptor and modulation of signal transduction pathways in human lens epithelial cells.

PURPOSE: Prolonged use of glucocorticoids (GCs) can lead to cataract formation. Lens GC responses have been difficult to elucidate. A previous study showed the presence of the glucocorticoid receptor (GR) in immortalized and primary human lens epithelial cells (hLECs) and GC-induced changes in gene expression. This study demonstrates specific GR activation and identifies the biological effect of GC-induced changes in gene expression in hLECs. METHODS: HLE B-3 (B-3) and primary cultures of hLECs were transfected with pGRE.Luc and treated with or without dexamethasone (Dex), RU-486, spironolactone, or vehicle. mRNA and protein expression were examined by real-time PCR and Western blot analysis, respectively. Cell proliferation and apoptosis were examined by WST-1 and flow cytometry, respectively. RESULTS: Dex treatment of B-3 and primary cultures demonstrated specific GR, but not mineralocorticoid receptor (MR), activation and phosphorylation. Pathway analysis revealed GC-induced changes in expression of MAPK regulators. Increased expression of GILZ mRNA and MKP-1 mRNA and protein was observed in immortalized and donor hLECs. This corresponded with a decrease in the phosphorylated forms of RAF, ERK, p38, and AKT, but not in JNK. No net change in LEC proliferation or apoptosis was observed with Dex treatment. CONCLUSIONS: GC treatment of hLECs activates the GR to modulate the expression of MAPK and PI3K/AKT regulators. This is the first demonstration of GC signaling in hLECs. GCs, MAPK, and PI3K/AKT are involved in cell processes implicated in steroid-induced cataractogenesis. The absence of a net change in cell activity with acute steroid treatment is consistent with the possibility that chronic treatment leads to prolonged modulation of these pathways and steroid-induced cataract.

Global gene profiling reveals novel glucocorticoid induced changes in gene expression of human lens epithelial cells.

PURPOSE: Prolonged use of glucocorticoids can lead to the formation of a cataract, however the mechanism is not known. We recently reported the presence of the functional glucocorticoid receptor in immortalized cultured mammalian lens epithelial cells (LECs), but the biological effect is not known. This study seeks to determine if freshly isolated human LECs respond to glucocorticoid treatment and to examine glucocorticoid induced changes in global gene expression in LECs. METHODS: Capsulorhexis specimens obtained in surgery from eyes with cataract were cultured. Primary lens cultures were transfected, in triplicate, with pGRE.Luc, which drives the expression of firefly luciferase, and treated with dexamethasone (Dex) or vehicle (Veh). RNA isolated from HLE B-3 cells, treated with Dex or Veh for 4 or 16 h in triplicate, was used to analyze global changes in gene expression by microarray hybridization. Data and cluster analyses were performed using Microarray Suite 5.0, GeneSpring 6.1, EASE, NetAffx, and SAM. Real Time PCR was used to confirm microarray data in RNA isolated from HLE B-3 cells in triplicate and a primary culture of human lens epithelial cells. RESULTS: Transfected primary cultures of human LECs treated with Dex demonstrated a glucocorticoid response with a greater than 4 fold increase in firefly luciferase activity over controls. Microarray data revealed that 136 genes were modulated with 4 h treatment with Dex. Of the 136 genes, 93 transcripts were upregulated and 43 were downregulated by greater than 1.5 fold. Eighty-six genes were modulated with 16 h Dex treatment. Of the 86 genes, 30 transcripts were upregulated and 56 were downregulated by greater than 1.5 fold. Microarray results were verified by Real Time PCR in both the HLE B-3 and primary cultures of lens epithelial cell. CONCLUSIONS: The activation of a GRE reporter gene in primary cultures of human LECs demonstrates that the glucocorticoid receptor is functional in non-immortalized human lens cells. Microarray studies at 2 time periods demonstrate that glucocorticoids modulate gene expression in immortalized human LECs, reveal novel changes in gene expression, and confirm an endogenous genomic lens glucocorticoid response. This study demonstrates that primary cultures of lens epithelial cells and microarray technology can be used to determine pathways involved in a lens glucocorticoid response and lead to a better understanding of the formation of a steroid induced cataract.

Association of polymorphisms in pulmonary surfactant protein A1 and A2 genes with high-altitude pulmonary edema.

STUDY OBJECTIVES: A potential pathogenetic cofactor for the development of high-altitude pulmonary edema (HAPE) is an increase in capillary permeability, which could occur as a result of an inflammatory reaction and/or free-radical-mediated injury to the lung. Pulmonary surfactant protein A (SP-A), the most abundant surfactant protein, has potent antioxidant properties and protects unsaturated phospholipids and growing cells from oxidative injury. Single-nucleotide polymorphisms (SNPs) in SP-A1 and SP-A2, genes encoding SP-A, have been associated with susceptibility to respiratory distress syndrome, COPD, and pulmonary infections. In view of the protective role of SP-A against inflammatory reactions and oxidative damage, the two underlying mechanisms in development of HAPE, we examined the association of constitutional susceptibility to HAPE with polymorphisms in SP-A1 and SP-A2. DESIGN: A cross-sectional case-control study. SETTING: Blood samples were collected at an altitude (> or = 3,500 m). PARTICIPANTS: Twelve low-altitude native (LAN) subjects with a history of HAPE, 15 healthy LAN sojourners without a history of HAPE (LAN control subjects), and 19 healthy high-altitude natives (HANs) without a history of HAPE (HAN control subjects). MEASUREMENTS: The SNPs in four exons and intermediate introns of the SP-A1 and SP-A2 were screened by polymerase chain reaction and sequencing. Biochemical parameters related to oxidative stress (malondialdehyde and reduced glutathione in RBC) and membrane permeability (circulating levels of lactate dehydrogenase) were measured in plasma. RESULTS: Allele frequencies of three loci in SP-A1 and one in SP-A2 were significantly different between LAN HAPE patients (SP-A1 C1101T: C allele, 36.4% and T allele, 63.6%; SP-A1 T3192C: T allele, 61.1% and C allele, 38.9%; SP-A1 T3234C: T allele, 61.1% and C allele, 38.9%; and SP-A2 A3265C: A allele, 21.4% and C allele, 78.6%) and LAN control subjects (SP-A1 C1101T: C allele, 8.3% and T allele, 91.7%; SP-A1 T3192C: T allele, 15% and C allele, 85%; SP-A1 T3234C: T allele, 15% and C allele, 85%; and SP-A2 A3265C: A allele, 37.5% and C allele, 62.5%) [C1101T odds ratio [OR], 6.3 with 95% confidence interval (CI), 2.8 to 14.3; T3192C OR, 8.9 with 95% CI, 4.5 to 17.6; T3234C OR, 8.9 with 95% CI, 4.5 to 17.6; and A3265C OR, 2.2 with 95% CI, 1.2 to 4.1 (p < or = 0.01)]. Heterozygous individuals, with respect to SP-A1 C1101T and SP-A2 A3265C, showed less severity in oxidative damage in comparison with homozygous subjects (SP-A1 T1101 and SP-A2 C3265). CONCLUSION: The polymorphisms in SP-A1 (C1101T, T3192C, and T3234C) and SP-A2 (A3265C) might be one of the genetic factors contributing to susceptibility to HAPE.

Expression of the functional glucocorticoid receptor in mouse and human lens epithelial cells.

PURPOSE: Studies have questioned the reported presence of a classic glucocorticoid receptor (GR) in the mammalian lens. The purpose of this study is to determine whether the functional GR is expressed in human and mouse lens epithelial cells. METHODS: GR mRNA was determined by RT-PCR in freshly isolated human lens epithelia, mouse lens, immortalized human (HLE B-3) and mouse (alphaTN4) lens epithelial cells and in mouse lung, NIH-3T3 cells, and HeLa cells, which served as positive controls. Western blot analysis with the GR-specific antibody H-300 was performed on protein extracts from human lens epithelia, HLE B-3 cells, and alphaTN4 cells and from HeLa cells, NIH-3T3 cells, and partially purified GR, which served as positive controls. pGRE.Luc drives the expression of firefly luciferase. HLE B-3 and alphaTN4 cells were transfected with pGRE.Luc and cotreated with dexamethasone, with and without the competitive inhibitor RU-486. RESULTS: PCR products of the expected size were detected in all samples, sequenced in both directions, and found to have 97% to 100% homology with the GR. A band in the appropriate molecular weight range was identified by Western blot analysis in the lens extracts. Active GR binding to the GRE was demonstrated by an increase in firefly luciferase expression in transfected cells treated with dexamethasone. The dexamethasone-induced increase in luciferase activity was inhibited with the addition of RU-486. CONCLUSIONS: These results demonstrate expression of the functional glucocorticoid receptor in mouse and human lens epithelial cells. This finding suggests that glucocorticoids may act on the mouse and human lens directly during normal lens development and/or cataractogenesis.

Antistress and adaptogenic activity of lecithin supplementation.

OBJECTIVE: Investigation of lecithin administration in rats for its antistress and adaptogenic activity. DESIGN: Lecithin was administered orally (1 mg/g body weight) to rats 0.5 hour, 1.5 hours, 3.0 hours, and 6.0 hours prior to cold (5 degrees C)-hypoxia (428 mm Hg)-restraint (C-H-R) exposure and serum choline estimation. The rectal temperature (T(rec)) of the rats was monitored during the exposure and the recovery periods. The time for decrease of T(rec) to 23 degrees C and its recovery to 37 degrees C were used as indices of antistress and adaptogenic activity. RESULTS: Lecithin administration 3-6 hours prior to C-H-R exposure was found to have significant adaptogenic activity that correlated with increased serum choline levels. Lecithin administration prior to C-H-R exposure increased the stress tolerance of the rats by delaying the time for decrease of T(rec) to 23 degrees C during C-H-R exposure. It also helped shorten poststress recovery as observed by decreased time to attain T(rec) of 37 degrees C from C-H-R-induced hypothermia (T(rec), 23 degrees C). CONCLUSIONS: The results suggested that lecithin supplementation acted as an antistress and adaptogenic functional food. It improved resistance during C-H-R exposure and enhanced recovery from hypothermia.