Effects of Bacillus sp. MR-1/2 and magnetite nanoparticles on yield improvement of rice by urea fertilizer under different watering regimes.

AIMS: The present research aimed to examine the use of magnetite nanoparticles (MNPs) in combination with phyto-beneficial rhizobacterium (PhBR) for improvement of applied N recovery (ANR) from urea fertilizer in rice grown under deficient and optimum watering conditions. METHODS AND RESULTS: The Bacillus sp. MR-1/2 was positive for acetylene reduction, phosphate solubilization and ACC deaminase activity at temperature ranges 35-45°C. In a pot experiment, urea, MNPs and Bacillus sp. MR-1/2 were applied either alone or in combination to rice plants grown in pots under water deficit and optimal watering conditions. Combined application of urea, MNPs and Bacillus sp. MR-1/2 increased the plant N content and ANR by 27 and 65%, respectively, over their respective control values in rice grown under optimum watering conditions, whereas these increases were 27 and 41%, respectively, in rice grown under water deficit conditions. This treatment also increased the kernel weight and plant dry matter by 36 and 60%, respectively, over control (urea alone) values in rice grown under water deficit conditions, whereas these increases were 31 and 21·8%, respectively, in rice grown under optimum watering conditions. Values of malondialdehyde (MDA) contents, ascorbate peroxidase (APX), catalase and ethylene concentration were higher in control treatment under both the watering regimes. The application of Bacillus sp. MR-1/2 either alone or in combination with MNPs and urea reduced MDA contents, APX, catalase and ethylene production in the rice plants. CONCLUSION: The combined application of MNPs+Bacillus sp. MR-1/2 reduced the N losses from applied urea, increased N uptake and ANR in rice, decreased MDA contents, APX and catalase activity and ethylene level in rice grown under deficit and optimum water conditions. SIGNIFICANCE AND IMPACT OF THE STUDY: The application of MNPs together with Bacillus sp. MR-1/2 may help to increase ANR and rice productivity under water deficit conditions with low cost of production.

Pathogenetic implication of fusion genes in acute promyelocytic leukemia and their diagnostic utility.

Acute promyelocytic leukemia (APL) has been recognized as a discrete subset of hematopoietic malignancies constituting approximately 10% of acute myeloid leukemia cases. The hallmark reciprocal chromosomal translocation t(15;17) involving fusion between the retinoic acid receptor (RARα) gene and promyelocytic leukemia (PML) gene is a characteristic feature in APL which consequently results in the emergence of PML-RARα chimeric gene. This gene has been substantiated to be responsible for cellular transformation and is a prime target of all-trans-retinoic acid (ATRA) as well as arsenic-trioxide (ATO) therapy. Since this initial discovery, about 10 diverse translocation partner genes of RARα have been reported that result in variant APL forms strongly suggesting that disruption of RARα underlies its pathogenesis. The nature of the fusion partner has a significant bearing upon disease characteristics including sensitivity to retinoids and ATO and thereby underpins the need for rapid and accurate diagnosis and also demands a highly specific treatment approach. In this article we laid emphasis on the rearrangement of the RARα gene and its different fusion partners resulting in variant forms of APL, their implication in underlying molecular pathogenesis of APL and also the different diagnostic modalities that should be employed for their rapid and accurate diagnosis.

Genetic investigation of 93 families with microphthalmia or posterior microphthalmos.

Microphthalmia is a developmental eye defect that is highly variable in severity and in its potential for systemic association. Despite the discovery of many disease genes in microphthalmia, at least 50% of patients remain undiagnosed genetically. Here, we describe a cohort of 147 patients (93 families) from our highly consanguineous population with various forms of microphthalmia (including the distinct entity of posterior microphthalmos) that were investigated using a next-generation sequencing multi-gene panel (i-panel) as well as whole exome sequencing and molecular karyotyping. A potentially causal mutation was identified in the majority of the cohort with microphthalmia (61%) and posterior microphthalmos (82%). The identified mutations (55 point mutations, 15 of which are novel) spanned 24 known disease genes, some of which have not or only very rarely been linked to microphthalmia (PAX6, SLC18A2, DSC3 and CNKSR1). Our study has also identified interesting candidate variants in 2 genes that have not been linked to human diseases (MYO10 and ZNF219), which we present here as novel candidates for microphthalmia. In addition to revealing novel phenotypic aspects of microphthalmia, this study expands its allelic and locus heterogeneity and highlights the need for expanded testing of patients with this condition.

IgE FcɛR1ß polymorphism and risk of developing chronic spontaneous urticaria: A study in an ethnic Kashmiri population.

BACKGROUND: The pathogenesis of chronic spontaneous urticaria involves interplay between the genetic and environmental factors, most of which is still poorly understood. It is well-recognized that 30-40% of chronic spontaneous urticaria is autoimmune in nature. Chronic autoimmune urticaria is caused by anti-FcɛR1ß and less frequently, by anti-IgE auto antibodies that lead to mast cell and basophil activation, thereby giving rise to the release of histamine and other proinflammatory mediators. We investigated the association between SNP loci in FcɛR1ß and chronic spontaneous urticaria and to see its relation with serum IgE levels in Kashmiri population. METHODS: The autologous serum skin test was used as a screening test for chronic autoimmune urticaria. PCR-RFLP was used to detect the genotype of the SNP loci. Serum IgE levels were assessed by ELISA kit. RESULTS: No significant difference was found between the study population and control group in genotype distribution (wild and variant) among FcɛR1ß loci (P value=0.06, odds ratio=0.29). The frequency of FcɛR1ß (C109T) in autologous serum skin test positive chronic autoimmune urticaria patients with the CT genotype was found to be statistically non-significant when compared with the wild genotype (P=0.35). Carriers of FcɛR1ß (T allele) had a more significant risk of developing CAU than those with C allele (P=0.01). In our population serum total IgE levels did not find any statistical significance with regard to ASST positive & ASST negative patients (P=0.26). CONCLUSIONS: There is statistically no significant association between FcɛR1ß gene polymorphism and CSU in Kashmiri population; however, there is a probability of developing CSU in patients carrying FcɛR1ß T allele. Furthermore, serum total IgE levels had no significant association with the development of CAU.

Genetic polymorphism of the glutathione-S-transferase P1 gene (GSTP1) and susceptibility to prostate cancer in the Kashmiri population.

Glutathione-S-transferase P1 (GSTP1) is a critical enzyme of the phase II detoxification pathway. One of the common functional polymorphisms of GSTP1 is A→G at nucleotide 313, which results in an amino acid substitution (Ile105Val) at the substrate binding site of GSTP1 and reduces catalytic activity of GSTP1. To investigate the GSTP1 Ile105Val genotype frequency in prostate cancer cases in the Kashmiri population, we designed a case-control study, in which 50 prostate cancer cases and 45 benign prostate hyperplasia cases were studied for GSTP1 Ile105Val polymorphism, compared to 80 controls taken from the general population, employing the PCR-RFLP technique. We found the frequency of the three different genotypes of GSTP1 Ile105Val in our ethnic Kashmir population, i.e., Ile/Ile, Ile/Val and Val/Val, to be 52.4, 33.3 and 14.3% among prostate cancer cases, 48.5, 37.5 and 14% among benign prostate hyperplasia cases and 73.8, 21.3 and 5% in the control population, respectively. There was a significant association between the GSTP1 Ile/Val genotype and the advanced age group among the cases. We conclude that GSTP1 Ile/Val polymorphism is involved in the risk of prostate cancer development in our population.

Risk of colorectal cancer associated with the methylenetetrahydrofolate reductase (MTHFR) C677T polymorphism in the Kashmiri population.

Methylenetetrahydrofolate reductase (MTHFR) is a critical enzyme in folate metabolism and is involved in DNA synthesis, DNA repair and DNA methylation. The two common functional polymorphisms of MTHFR, 677 C→T and 1298 A→C, have been shown to impact various diseases, including cancer. The 677 C→T polymorphism has been widely investigated in different cancers and has been implicated as a risk factor for the development of various cancers. We investigated MTHFR C677T genotype frequency in colorectal cancer cases in the Kashmiri population and correlated this information with the known clinicopathological characters of colorectal cancer, in a case-control study. Eighty-six colorectal cancer cases were studied for MTHFR C677T polymorphism, compared to 160 controls taken from the general population, employing the PCR-RFLP technique. We found the frequency of the three different genotypes of MTHFR in our ethnic Kashmir population, i.e., CC, CT and TT, to be 68.6, 20.9 and 10.4% among colorectal cancer cases and 75.6, 16.9 and 7.5% among the general control population, respectively. There was a significant association between the MTHFR TT genotype and colorectal cancer in the higher age group. We conclude that the MTHFR C677T polymorphism slightly increases the risk for colorectal cancer development in our ethnic Kashmir population.

Heme oxygenase 1, beneficial role in permanent ischemic stroke and in Gingko biloba (EGb 761) neuroprotection.

Ginkgo biloba extract, EGb 761, a popular and standardized natural extract, contains 24% ginkgo-flavonol glycosides and 6% terpene lactones. EGb 761 is used worldwide to treat many ailments, and although a number of studies have shown its neuroprotective properties, the mechanisms of action have not been elucidated fully. We hypothesize that EGb 761 and some of its bioactive components [Bilobalide (BB), Ginkgolide A (GA), Ginkgolide B (GB), and Terpene Free Material (TFM)] could provide neuroprotection in ischemic conditions through heme oxygenase 1 (HO1). Mice were subjected to permanent distal middle cerebral artery occlusion (pMCAO) and survived for 7 days. HO1 knockout (HO1⁻/⁻) mice showed significantly higher (P<0.05) infarct volume and Neurologic Deficit Scores (NDS) as compared to their wildtype (WT) counterparts. In another cohort, WT mice subjected to pMCAO and treated at 4 h of pMCAO with 100 mg/kg EGb 761, 6 mg/kg BB, GA, GB, or 10 mg/kg TFM showed significantly lower (P<0.05) infarct volumes (BB; 29.0±3.9%, GA; 31.3±4.0%, GB; 32.0±3.8%, TFM; 32.5±3.5%, and EGb 761; 27.4±4.5%) than those in the vehicle-treated mice (46.0±3.7%). Similarly, NDS were lower in BB; 7.1±1.8, GA; 7.4±2.1, GB; 7.9±1.8, TFM; 7.7±1.7, and EGb 761; 6.8±2.0 groups as compared with the vehicle-treated group (13.8±1.5). Interestingly, the protective effect of EGb 761 was essentially lost when HO1 knockout mice were treated with EGb 761. In another cohort, HO1, vascular endothelial growth factor (VEGF) and endothelial nitric oxide synthase (eNOS) protein levels in the brain cortices appeared to be higher in EGb 761 and BB but not in GA, GB and TFM treated groups. Together, these results suggest that HO1 plays, at least in part, an important role in the neuroprotective mechanism of EGb 761 and in delayed ischemia. Targeting this pathway could lead to neuroprotective agents against ischemic stroke.

Neuroprotective properties of prostaglandin I2 IP receptor in focal cerebral ischemia.

We and others have identified that inhibition of cyclooxygenase might not be the optimal approach to limiting brain damage after stroke. Now we are investigating the unique properties of the various prostaglandin receptors to determine whether blocking those that mediate toxicity or stimulating those that reduce toxicity will improve neurological outcomes. Here, we determined the respective contribution of the prostaglandin I(2) (PGI(2)) receptor in transient middle cerebral artery (MCA) occlusion (tMCAO) and permanent MCAO (pMCAO) preclinical stroke models by using male wildtype (WT) and IP receptor knockout (IP(-/-)) C57Bl/6 mice. In addition, we investigated the putative preventive and therapeutic effects of the IP receptor agonist beraprost. The infarct volumes and neurological deficit scores (NDS) were significantly greater in IP(-/-) than in WT mice after both tMCAO and pMCAO. Interestingly, beraprost pretreatment (50 or 100 microg/kg p.o.) 30 min before tMCAO and post-treatment (100 microg/kg p.o.) at 2 or 4.5 h of reperfusion significantly reduced the neurological deficit score and infarct volume in WT mice. Post-treatment with beraprost (100 microg/kg p.o.) 4.5 h after pMCAO also significantly decreased neurological deficits and infarct volume in WT mice. Together, these novel findings suggest for the first time that PGI(2) IP receptor activation can attenuate anatomical and functional damage following ischemic stroke.

TP53 Pro47Ser and Arg72Pro polymorphisms and colorectal cancer predisposition in an ethnic Kashmiri population.

Two TP53 gene polymorphisms at codon 47 (TP53 Pro47Ser) and at codon 72 (TP53 Arg72Pro) have been associated with susceptibility to various cancers. We carried out a case-control study and examined the genotype distribution of TP53 Pro47Ser and Arg72Pro single nucleotide polymorphisms (SNPs), using a PCR-RFLP approach, to determine if these two SNPs are risk factors for colorectal cancer (CRC) development and to look for a possible correlation of these two SNPs with clinicopathological variables of CRC. We investigated the genotype distribution of these SNPs in 86 CRC cases in comparison with 160 healthy subjects in an ethnic Kashmiri population. TP53 Arg72Pro SNP genotype frequencies differed significantly (P = 0.000001) between the groups; the frequency of the Pro/Pro mutant was almost 20% in the general population. We also found significant association of the Pro/Pro mutant with tumor location, nodal status/higher tumor grade and bleeding per rectum/constipation. We conclude that Arg72Pro SNP is associated with susceptibility to developing CRC in this ethnic Kashmiri population.

Mutation pattern of K-ras gene in colorectal cancer patients of Kashmir: a report.

BACKGROUND AND AIM: Colorectal cancer (CRC) is one of the leading malignancies worldwide. CRC has been reported to show geographical variation in its incidence, even within areas of ethnic homogeneity. The aim of this study is to identify K-ras gene mutations in CRC patients among the Kashmiri population, and to assess whether they are linked with the clinicopathological parameters. MATERIALS AND METHODS: Paired tumor and normal tissue samples were collected from a consecutive series of 53 patients undergoing resective surgery for CRC. In addition blood was also collected from all the cases for ruling out germline mutation. RESULTS: Colorectal patients, 22.64% (12 of 53), presented with mutations in K-ras constituting 13 missense mutations out of which 11 were G-->A transition, one G-->C transversion, and one G-->T transversion. 61.5% percent of the mutations occurred in codon 12 and 38.5% in codon 13. One tumor contained missense mutations in both codons. K-ras mutations were significantly associated with advanced Dukes' stage (P < 0.05) and positive lymph node status (P < 0.05). Moreover Codon 12 K-ras mutations were associated with mucinous histotype (P < 0.05). Comparison of the mutation profile with other high-risk areas reflected both mucinous histotype differences and similarities indicating coexposure to a unique set of risk factors. CONCLUSION: Mutation of the K-ras gene is one of the commonest genetic changes in the development of human CRC, but it occurs in a rather low frequency in Kashmiri population.

Lipocalin-prostaglandin D synthase is a critical beneficial factor in transient and permanent focal cerebral ischemia.

Prostaglandin D(2) (PGD(2)) is the most abundant prostaglandin produced in the brain. It is a metabolite of arachidonic acid and synthesized by prostaglandin D(2) synthases (PGDS) via the cyclooxygenase pathway. Two distinct types of PGDS have been identified: hematopoietic prostaglandin D synthase (H-PGDS) and lipocalin-type prostaglandin D synthase (L-PGDS). Because relatively little is known about the role of L-PGDS in the CNS, here we examined the outcomes in L-PGDS knockout and wild-type (WT) mice after two different cerebral ischemia models, transient middle cerebral artery (MCA) occlusion (tMCAO) and permanent distal middle cerebral artery occlusion (pMCAO). In the tMCAO model, the MCA was occluded with a monofilament for 90 min and then reperfused for 4 days. In the pMCAO model, the distal part of the MCA was permanently occluded and the mice were sacrificed after 7 days. Percent corrected infarct volume and neurological score were determined after 4 and 7 days, respectively. L-PGDS knockout mice had significantly greater infarct volume and brain edema than did WT mice after tMCAO (P<0.01). Similarly, L-PGDS knockout mice showed greater infarct volume and neurological deficits as compared to their WT counterparts after pMCAO (P<0.01). Using the two models enabled us to study the role of L-PGDS in both early (tMCAO) and delayed (pMCAO) ischemic processes. Our findings suggest that L-PGDS is beneficial for protecting the brain against transient and permanent cerebral ischemia. These results provide a better understanding of the role played by the enzymes that control eicosanoid synthesis and how they can be utilized as potential targets to prevent damage following either acute or potentially chronic neurological disorders.

Vitamin D status in apparently healthy adults in Kashmir Valley of Indian subcontinent.

BACKGROUND: The worldwide prevalence of vitamin D deficiency is reported to be high. OBJECTIVES: To assess the vitamin D status in apparently healthy adults in Kashmir valley by measuring serum 25-hydroxyvitamin D (25 (OH) D). METHODS: 92 healthy natives (64 men and 28 non-pregnant/non-lactating women, aged 18-40 years), residing in Kashmir for at least last 5 years and not having any suggestion of systemic disease, were selected for this study. The samples were collected throughout the year in both summer and winter months. Vitamin D deficiency was defined as a serum 25 (OH) D concentration of <50 nmol/l and graded as mild (25-50 nmol/l), moderate (12.5-25 nmol/l) and severe (<12.5 nmol/l). RESULTS: Body mass index, total energy intake, and other nutritional parameters were comparable among subjects in different groups. Overall 76 (83%) of the subjects studied had vitamin D deficiency--25%, 33%, and 25% had mild, moderate, and severe deficiency, respectively. 49 of the 64 males and all but 1 of the 28 females were vitamin D deficient. The prevalence of vitamin D deficiency ranged from 69.6% in the employed group to 100% in the household group. Vitamin D deficiency was equally prevalent in subjects from rural and urban areas. Serum calcium and phosphorus values were comparable in subjects with and without vitamin D deficiency, while daily intake and urinary excretion of calcium were significantly lower in the former. Vitamin D deficient subjects had a significantly lower mean weekly exposure to sunlight. CONCLUSIONS: In spite of abundant sunlight, healthy individuals in Kashmir valley are vitamin D deficient, particularly women. Serum 25 (OH) D concentrations are significantly related to sun exposure.

Role of reactive oxygen species in modulation of Nrf2 following ischemic reperfusion injury.

The transcriptional factor Nrf2 has a unique role in various physiological stress conditions, but its contribution to ischemia/reperfusion injury has not been fully explored. Therefore, wildtype (WT) and Nrf2 knockout (Nrf2(-/-)) mice were subjected to 90-min occlusion of the middle cerebral artery (MCA) followed by 24-h reperfusion to elucidate Nrf2 contribution in protecting against ischemia/reperfusion injury. Infarct volume, represented as percent of hemispheric volume, was significantly (P<0.05) larger in Nrf2(-/-) mice than in WT mice (30.8+/-6.1 vs. 17.0+/-5.1%). Furthermore, neurological deficit was significantly greater in the Nrf2(-/-) mice. To examine whether neuronal protection was mediated by Nrf2, neurons were treated with various compounds to induce excitotoxic or oxidative stress. Translocation of Nrf2 into the nucleus was increased by the free-radical donor tert-butylhydroperoxide, but not by glutamate or N-methyl-D-aspartic acid (NMDA). In addition, a common Nrf2 inducer, tert-butylhydroquinone, significantly attenuated neuronal cell death induced by tert-butylhydroperoxide (83.6+/-1.6 vs. 62.0+/-7.7%) but not as substantially when excitotoxicity was induced by NMDA (91.9+/-1.6 vs. 79.3+/-3.3%) or glutamate (87.8+/-1.5 vs. 80.2+/-2.6%). The results suggest that Nrf2 reduces ischemic brain injury by protecting against oxidative stress.

An immunological based study of monoclonal gammopathies among suspected individuals in Kashmir region.

Serum and urine samples from 513 patients clinically suspected of monoclonal gammopathies over a period of five years (1992-97) were subjected to various immunological procedures viz, electrophoresis, immunoelectrophoresis and immunoglobulin estimations. Laboratory investigations confirmed gammopathies in 10.33%. It was observed that overall age of incidence for monoclonal gammopathies in both sexes was between 42-72 years with a male to female ratio of 1.4∶1. Predominant paraprotein detected was IgG type (75.47%) followed by IgA (16.98%) and Bence Jones proteins (7.55%). Amongst positive patients, 64.16% were having kappa (k) type light chains and 35.84% lambda (δ) type light chains. 69.39% patients with serum M component (IgG and IgA) had Bence Jones proteinuria. Densitometric scanning revealed that majority of IgG type paraprotein was found in the slow gamma globulin region and majority of IgA type paraprotein was found equally distributed between beta and fast gamma globulin regions. Both types had decreased albumin and alpha-2-globulin concentrations as compared to normal controls. Immunoglobulin levels in patients with paraprotein had very high levels of serum IgG (6467.0 mg%) and IgA (2714.0 mg%) in respective types of monoclonal gammopathies; the rest of immunoglobulin classes were either at normal or decreased levels.