Association of pro/anti-inflammatory cytokine gene variants in renal transplant patients with allograft outcome and cyclosporine immunosuppressant levels

T-helper (Th) type 1/Th2 cytokines are key mediators in induction/effecter phases of all immune and inflammatory responses playing role in acute/chronic renal allograft rejection. Association studies lead to identification of patient risk profiles enabling individualization of level of immunosuppressions. We investigated the association of allograft rejection with interleukin-2 (IL-2), IL-4, IL-6, tumor necrosis factor-α (TNF-α) −308, transforming growth factor-β (TGF-β) (C-del, codon 10 and 25) gene variants in 184 renal transplant recipients and 180 controls. These cytokine genotypes were also evaluated with cyclosporine levels (C2) at one month in 135 stable recipients. High producing genotypes B1B1 of IL-4 and AA of TNF-α α308 showed significant association with rejection of allograft. The dose-adjusted C2 levels were significantly lower in patients with the high producing genotype T/T of IL-2 and heterozygous G/C of TGF-β codon 25 (P = 0.012 and 0.010, respectively). Haplotype frequencies were comparable in subjects for TGF-β codon-10 and 25. Combined inter-gene interaction showed high risk for rejection in recipients with high producing genotype B1B1 of IL-4 and AA of TNF-α and high TNF-α (AA) with low TGF-β (CC or Pro/Pro). In conclusion, association of IL-4 VNTR and TNF-α −308 suggested the involvement of these cytokines contributing to pathogenesis of allograft rejection. Recipients with TT genotype of IL-2 and GC of TGF-β codon 25 having low C2 levels may require higher cyclosporine dosage. Combined analysis of gene-gene interaction demonstrated synergistic effect of cytokines increasing risk for rejection. Thus, this information may help in pre-assessment of allograft outcome and to optimize cyclosporine therapy in post-transplant patients

Amelioration of experimental rheumatoid arthritis by selected ultra-diluted preparations by down regulating increased expression of TNF-α & IL-6

956-964The current work explored the inhibitory effect of selected homeopathic drugs, in experimental models of inflammation and CFA-induced arthritis. Twelve groups of animals were made, each containing 6 animals. The selected homeopathic drugs (causticum, calcarea, medorrhinum, mercurius, formica, proteus, silica, sulphur, thuja), placebo and standard drug, Indomethacin. In CFA model, treatment groups and the reference drug were administered daily for a period of 21 days. Dysfunction in joints was evaluated by parameters such as joint diameter, expression of inflammatory markers (TNF-α, IL-6). Findings of the study revealed that on CFA administration, there is a significant (p<0.01) increase in joint diameter in all the tested animals. On day 3, we found highest increase in the joint diameter in all treatment groups. Medorrhinum, silica, sulphur showed significant (p<0.01) decrease in joint diameter on day 21. Significant (p<0.05) reduction in paw edema was observed at 5 h post carrageenan administration. IHC of NF-kB in CFA treated group revealed presence of vacuoles, infiltration of inflammatory cells. However, prominent reversal of joint damage was seen in homeopathic drugs (medorrhinum, silica, sulphur) and indomethacin. Study inferred that the homeopathic drugs (medorrhinum, silica, sulphur) and indomethacin were found to be potent in ameliorating inflammation

PRIMARY CELL CULTURE OF AEDES ALBOPICTUS MIDGUT CELLS: A PROSPECTIVE MODEL FOR IN VITRO STUDY OF ARBOVIRUSES

  Objective: Midgut cells play a key role in the propagation of mosquito borne Arboviruses. The existing mosquito cell lines for studying viral pathogenesis are derived either from larvae or from eggs since there is no cell line available from the mosquito midgut. Therefore, to delineate the in situ viral interaction which naturally occurs within the mosquito midgut and represent cellular pathogenesis in human beings, the present work was aimed to develop a primary cell line from the midgut cells of Aedes albopictus.Methods: The midgut cells of A. albopictus were collected, cultured and incubated at 28°C to study the growth after every 24 hrs for 7 days.Result: The primary cell culture showed an increasing growth pattern of columnar cells up to 48 hrs followed by decrease in cell population afterward. However, the number of stem cells increased significantly throughout the study period, and their population outnumbered the columnar cells after 72 hrs. There was no significant change of goblet cells and regenerating cells which were scanty in number throughout the experiment.Conclusion: The present method will help to develop the individual cell lines from mosquito midgut and study the host pathogen interaction in arboviral diseases in future

Assessment of the effectiveness of homoeopathic remedies in improving quality of life of chronic urticaria patients in a typical clinical setting

Objective: To evaluate the effectiveness of homoeopathic remedies in improving quality of life (QoL) of chronic urticaria (CU) patients. Methods: Setting: The study population included patients attending the Outpatient Department of State Homoeopathic Dispensary, Ahmadpur, Aligarh, Uttar Pradesh, India. CU-QoL questionnaire (CU-Q2oL) and average Urticaria Activity Score for 7 days (UAS7) questionnaires were filled questionnaires were filled at baseline and 3rd, 6th, 9th and 12th months. The study included both male and female patients clinically diagnosed with CU, screened from January 2015 to June 2016. Eighteen homoeopathic remedies were used. The individualised prescription was based on the totality of each patient's symptoms. Scores were analysed using one-way repeated measures ANOVA with SPSS version 19. Results: A total of 134 patients were screened and 70 were diagnosed with CU and enrolled in the study were analysed under modified intention-to-treat approach. Significant difference was found in baseline and 12th month CU-Q2oL score (mean difference 34.14 with standard error of 1.65, 95% confidence interval, lower bound 29.31, upper limit 38.94, P < 0.001). A one-way repeated measures ANOVA was calculated for comparing CU-Q2oL scores (F [2.45, 169.46] = 260.89, P ≤ 0.000, effect size = 0.791). Apis mellifica (n = 10), Natrum muriaticum (n = 9), Rhus toxicodendron (n = 8) and Sulphur (n = 8) were the most frequently used remedies. Conclusions: Homoeopathic medicines have potential to improve QoL of CU patients by reducing pruritus, intensity of wheals, swelling, nervousness, and improve sleep, mood and concentration. Further studies with more sample size are desirable

Comparative standardization study for determination of reserpine in Rauwolfia serpentina homoeopathic mother tinctures manufactured by different pharmaceutical industries using HPTLC as a check for quality control

Background: Rauwolfia serpentina (L.) Benth. ex Kurz (Apocynaceae) (Indian snakeroot), popularly known as Sarpagandha (Sanskrit), is used for the treatment of insanity, fever, snake bites, anxiety and in neuropsychiatric conditions. The antihypertensive actions of Reserpine are a result of its ability to deplete catecholamines (amongst other monoamine neurotransmitters) from peripheral sympathetic nerve endings which are normally involved in controlling heart rate, force of cardiac contraction and peripheral vascular resistance. Objective: Comparative study of Reserpine content in R. serpentina homoeopathic mother tinctures manufactured by different pharmaceutical industries and in-house mother tinctures applying high-performance thin-layer chromatography investigative techniques to facilitate the use of correct species. Materials and Methods: The authentic samples of roots of R. serpentina were supplied by Centre of Medicinal Plants Research in Homoeopathy, Emerald, Tamil Nadu, India. Authentic plant material was used to prepare the mother tincture (as per Homoeopathic Pharmacopoeia of India). Reserpine (C33H40N2O9,M.P. 360°C, purity >99% w/w by high-performance liquid chromatography [HPLC]) was purchased from Sigma-Aldrich as a standard reference. The solvents for the study, namely, ethanol, HPLC water, toluene, ethyl acetate, diethylamine and chloroform were of analytical grade purity (MERCK Ltd.,), used throughout. Results: Five samples of mother tinctures were used for the study, in-house mother tinctures (labelled: D and E) of R. serpentina shows a higher amount of Reserpine content than the marketed samples (labelled: A, B and C). Conclusion: It may be concluded that mother tinctures prepared by authentic plants showed the excess amount of Reserpine rather than that of mother tinctures procured from the market

Safety studies of homoeopathic drugs in acute, sub-acute and chronic toxicity in rats

Background: Homoeopathic drugs are frequently recommended in day to day life as therapeutic agents by homoeopathic practitioners. However, safety of homoeopathic drugs remains a challenge because of the high variability of chemical components involved. Aim: The objective of the present study was to investigate the acute, subacute, and chronic oral toxicity of different homoeopathic drugs (Ferrum phosphoricum 3X, Ferrum phosphoricum 6X, Calcarea phosphoricum 6X, and Magnesium phosphoricum 6X) in experimental models. Materials and Methods: In acute oral toxicity study, homoeopathic drugs were administered orally at 2000mg/kg body weight, and animals were observed for toxic symptoms till 10 days as per the OECD guidelines. For subacute and chronic toxicity study, homoeopathic drugs were administered for 28 and 180 days, respectively, as per the OECD guidelines. At the end of 28 and 180 days, the animals were sacrificed and toxicity parameters were assessed. Histopathological evaluation of different organs was also performed to assess any toxicity. Results: In acute toxicity study, no mortality was found at a dose of 2000 mg/kg which indicates that oral LD50of homoeopathic drugs were more than 2000 mg/kg. The administration of drugs at a dose of 70 mg/kg body weight for 28 and 180 days did not produce any significant change in haematological and biochemical parameters of male and female rats as compared to normal control group. No pathological changes were observed in histology of various organs of treated rats as compared to normal control animals. Conclusion: These homoeopathic drugs are safe & produce no toxicity when administered for longer duration

Pharmacognostic study of Chamaecyparis lawsoniana (Murr.) Parl.: A drug used in Homoeopathy

The pharmacognostic profile of crude drug has a key role in standardization for quality, purity and drug identification. The present study deals with pharmacognostic evaluation of aerial part of Chamaecyparis lawsoniana (Murr.) Parl. a drug used in homoeopathic system of medicine for diverse clinical uses such as terrible pain in stomach, tumors, keloid, warts and lipoma of thigh. The study includes collection, identification, macroscopy, microscopy and organoleptic characteristics of aerial part of Chamaecyparis lawsoniana. Anatomically the leaf is distinguishable into a layer of the epidermis followed by parenchymatous mesophyll and resin duct in the parenchymatous cortex. Powder microscopy shows the presence of epidermal cells, parenchymatous cells and tracheids. These observations may be used as pharmacopoeial standards for identification of Cha maecyparis lawsoniana

An observation on direct changes in Aedes albopictus midgut cells by Rhus tox 6C in relation to dengue virus infection

Background and Objectives: In mosquito vectors, dengue virus (DENV) invasion occurs through midgut cells, but available mosquito cell lines for in vitro study of DENV are prepared from eggs or larvae, which are not appropriate models, to study its infectivity. Hence, we developed a new primary cell culture, from Aedes albopictus mosquito midgut, and standardized it for in vitro study of DENV, with an aim to find out any possible role of homoeopathic medicines, in preventing or reducing DENV invasiveness in these midgut cells. This midgut primary cell culture demonstrated prominent cytopathic effects on infection with wild DENV isolated from dengue-infected patients in viremic phase. Materials and Methods: In this paper, we observed the direct effect of homoeopathic medicine Rhus toxicodendron 6C (Rhus tox 6C) (ultra dilution of 10−12 ) on this primary cell culture, to find out significant changes, to be used as baseline data in future experiments to observe possible role of Rhus tox 6C against DENV infection in these cells. Hence, these direct changes may be a prerequisite for the action of this medicine against DENV invasion; as this is one of common repertoire homoeopathic medicines used against dengue fever. Conclusion and Discussion: In our experiments, we found that Rhus tox 6C could increase cell size and help organization of cells on the solid surface as observed under scanning electron microscope although the total number of cells was decreased. Moreover, Rhus tox 6C treated cells were healthier as indicated by less number of deformed, clump, and diploform cells

Chemoprofiling of homoeopathic drug Holarrhena antidysenterica L.

Background: Chemoprofiling of homoeopathic drug/tincture (HT) represents a comprehensive approach for evaluation of quality, purity, safety and efficacy of HT. This paper reflects the chemoprofiling of Homoeopathic drug Holarrhena antidysentrica L. Objective: The objective of this study is to standardise Holarrhena antidysenterica mother tincture by taking the samples from four different sources: Dr D. P. Rastogi, CRI (H) Noida (A) and three from market (labelled as B, C and D). Materials and Methods: The authentic sample of bark of Holarrhena antidysenterica supplied by the Centre of Medicinal Plants Research in Homoeopathy, Emerald, Tamil Nadu, India was used to prepare the mother tincture (as per the Homoeopathic Pharmacopoeia of India). The solvents used throughout the study, namely, ethanol, high-pressure liquid chromatography water, cyclohexane, chloromethane and diethylamine, were of analytical grade purity (MERCK Ltd.). Physicochemical properties, ultraviolet (UV) spectroscopy and high-performance thin-layer chromatography (HPTLC) chemoprofile of raw drug and mother tinctures were standardised and compared with market samples. Results: The present study reveals the moisture content (14.40%), total ash (4.65%), alcohol (18.0%), water extractive values (16.0%), total solids (1.47%), weight/ml (0.92 g) and alcohol content (60.6%). In UV spectroscopy, λmaxvalues were observed at 228 and 278 nm in HT. HPTLC analysis of in-house HT (A) and three market samples (B, C, D) was performed by using cyclohexane: chloromethane: diethylamine (7:3:1, v/v/v) as mobile phase. Under UV light (254, 366 nm) and in the presence of visualising agent Dragendroff, bands of active constituent were observed in all the four samples. However, excess amount of active constituents were found in in-house HT (a) rather than the market samples (B, C and D). Conclusion: The present physicochemical and phytochemical data may be considered as pharmacopoeia standards for the homoeopathic drug Holarrhema antidysentrica L

A multicentric, double-blind randomized, homoeopathic pathogenetic trial of Allium sativum

Background: Homoeopathic drug proving is an integral part of Homoeopathic System of Medicine. It is the first step in finding out the pathogenetic powers of a drug. Objective: To elicit the pathogenetic response to Allium sativum in homoeopathic potencies on healthy human provers. Materials and Methods: A multi-center randomized, placebo-controlled, double-blind trial was conducted at two centers of the Central Council for Research in Homoeopathy (CCRH). Proving was conducted on 33 healthy provers after the pretrial medical examination. All the provers were given 12 doses of placebo divided in 4 doses/day for 3 days during the first phase of the trial. After randomization, in the intervention group (21 provers), Allium sativum (A. sativum) was proved in 6C and 30C potencies, in two phases. In the placebo group, 12 provers were administered placebo in the same manner. The symptoms manifested during the trial period were noted down by the provers and then elaborated by the proving masters. The generated data on A. sativum were then compiled and analyzed at proving-cum-data processing cell at CCRH headquarters. Results: Out of 21 provers who were on actual drug trial, only nine provers manifested symptoms. Drug was able to manifest symptoms in both the potencies, in more or less every part of the body. Conclusion: The pathogenetic response elicited during the proving trial expands the scope of use of the drug A. sativum and will benefit the research scholars and clinicians. The generated symptoms of this drug will carry more value when verified clinically