Anticandidal activity of ethanolic root extract of Juglans regia (L.): Effect on growth, cell morphology, and key virulence factors.

The emergence of drug-resistant strains has encouraged several studies on natural products with antifungal activity and low toxicity. In this study, the antifungal effect of methanolic root extract of Juglans regia (JRE) was investigated against 9 strains of Candida (one reference and 8 clinical strains) through MIC90 and spot assays. To gain insight into the mechanism of antifungal action, we carried out confocal scanning laser microscopy (CLSM), transmission electron microscopy (TEM), and then examined the effect of JRE on hydrolytic enzyme secretion. Additionally, JRE was subjected to various phytochemical tests, chemically characterized by gas chromatography-mass spectrometry analysis (GC-MS) and its toxicity was tested against H9c2 rat cardiac myoblasts. The phytochemical tests showed the presence of phenols, alkaloids, steroids, saponins, and tannins in JRE. In the GC-MS analysis, a total of 40 compounds were identified. JRE was found to be effective in liquid media with MICs ranging from 300 to 700µg/mL. Spot assay results revealed that Candida cells show increased sensitivity to JRE. CSLM experiments showed that cells exposed to JRE (MIC) exhibited cell membrane disruption. TEM micrograph of treated cells showed extensive breakage in the cell wall and cell membrane. Average inhibition of proteinase and phospholipase secretion (of five C. albicans strains) at MIC/2 values of JRE was 45.17%, and 34.29%, respectively. Cellular toxicity of JRE against H9c2 rat cardiac myoblasts was less than 10% at the highest MIC value. These findings encourage further development of JRE.

Cerebrospinal fluid procalcitonin as a biomarker of bacterial meningitis in neonates.

OBJECTIVE: The objective of the study was to study the performance of cerebrospinal fluid (CSF) procalcitonin as a marker for bacterial meningitis in neonates, and to determine its optimal 'cutoff' in CSF that can be called significant for the diagnosis. STUDY DESIGN: Neonates qualifying for lumbar puncture were prospectively studied. Procalcitonin and established CSF parameters were recorded. RESULTS: At a cut-off value of 0.33 ng ml-1, CSF procalcitonin had a sensitivity of 0.92, specificity of 0.87, with positive and negative likelihood ratios of 7.13 and 0.092, respectively. The area under the curve for different CSF parameters was: 0.926 (0.887 to 0.964) (P<0.001) for procalcitonin, 0.965 (0.956 to 0.974) for total leukocyte count, 0.961 (0.94 to 0.983) for neutrophil count, 0.874 (0.825 to 0.923) for protein, 0.946 (0.914 to 0.978) for sugar and 0.92 (0.955 to 0.992) for CSF:serum sugar ratio. The lumbar puncture was traumatic in 36 (21.4%) patients; out of these 15 (41.7%) had bacterial meningitis and 21 (58.3%) had no meningitis. In traumatic lumbar tap group, the median (IQR) CSF procalcitonin in patients with and without meningitis was 1.41 (0.32-3.42) ng/ml and 0.21(0.20-0.31) ng/ml respectively (p<0.05). CONCLUSIONS: Procalcitonin measurement has diagnostic efficiency similar to the established CSF markers. Routine assessment of procalcitonin in clean non-contaminated CSF may not yield additional information, but it may have clinical utility in situations where diagnosis of meningitis is in dilemma, as in the case of blood contamination of CSF in traumatic lumbar punctures.

Quercetin attenuates neuronal death against aluminum-induced neurodegeneration in the rat hippocampus.

Aluminum is a light weight and toxic metal present ubiquitously on earth, which has gained considerable attention due to its neurotoxic effects. It also has been linked ecologically and epidemiologically to several neurological disorders, including Alzheimer's disease (AD), Parkinson's disease (PD), Guamanian-Parkinsonian complex and Amyotrophic lateral sclerosis (ALS). The mechanism of aluminum neurotoxicity is poorly understood, but it is well documented that aluminum generates reactive oxygen species (ROS). Enhanced ROS production leads to disruption of cellular antioxidant defense systems and release of cytochrome c (cyt-c) from mitochondria to cytosol resulting in apoptotic cell death. Quercetin (a natural flavonoid) protects it from oxidative damage and has been shown to decrease mitochondrial damage in various animal models of oxidative stress. We hypothesized that if oxidative damage to mitochondria does play a significant role in aluminum-induced neurodegeneration, and then quercetin should ameliorate neuronal apoptosis. Administration of quercetin (10 mg/kg body wt/day) reduced aluminum (10 mg/kg body wt/day)-induced oxidative stress (decreased ROS production, increased mitochondrial superoxide dismutase (MnSOD) activity). In addition, quercetin also prevents aluminum-induced translocation of cyt-c, and up-regulates Bcl-2, down-regulates Bax, p53, caspase-3 activation and reduces DNA fragmentation. Quercetin also obstructs aluminum-induced neurodegenerative changes in aluminum-treated rats as seen by Hematoxylin and Eosin (H&E) staining. Further electron microscopic studies revealed that quercetin attenuates aluminum-induced mitochondrial swelling, loss of cristae and chromatin condensation. These results indicate that treatment with quercetin may represent a therapeutic strategy to attenuate the neuronal death against aluminum-induced neurodegeneration.

Caspase inhibition augments Dichlorvos-induced dopaminergic neuronal cell death by increasing ROS production and PARP1 activation.

Numerous epidemiological studies have shown an association between pesticide exposure and the increased risk of developing Parkinson's disease. Previously we have reported that Dichlorvos exposure can induce oxidative stress, resulting in over-expression of pro-apoptotic genes and finally caspase-dependent nigrostriatal dopaminergic neuronal cell death in rat brain. Here, we examined the effect of caspase inhibition on PC12 cell death induced by Dichlorvos (30 µM). Reactive oxygen species (ROS) generation followed by protein carbonylation, lipid peroxidation, decreased antioxidant defenses (decreased Mn-superoxide dismutase (MnSOD) activity and decreased glutathione levels) and subsequent caspase activation mediated the apoptosis. Inhibition of caspase cascade with Boc-aspartyl(OMe)-fluoromethylketone (BAF) enhanced the Dichlorvos-induced PC12 cell death, as assessed by the increased cellular efflux of lactate dehydrogenase (LDH). This increase in cell death was accompanied by a marked increase in poly(ADP-ribose) polymerase-1 (PARP1) activity, increased oxidative stress, a reduction in the mitochondrial membrane potential and reduced cellular NAD and ATP levels. Pretreatment of cells with PJ34, a PARP1 inhibitor prevented the cells from undergoing cell death and preserved intracellular NAD and ATP levels. Subsequent release of the apoptosis-inducing factor (AIF) from mitochondria and its translocation into the nucleus was also prevented by PJ34 pretreatment. In conclusion, the results of the present study show that caspase inhibition without concurrent inhibition of PARP1 is unlikely to be effective in preventing cell death because in the presence of the caspase inhibitor, caspase-independent cell death predominates due to PARP activation. These results suggest that combined therapeutic strategies directed at multiple cell death pathways may provide superior neuroprotection than those directed at a single mechanism.

Protective efficacy of coenzyme Q10 against DDVP-induced cognitive impairments and neurodegeneration in rats.

The present study was carried out to elucidate the effects of coenzyme Q(10) (CoQ(10)) against cognitive impairments induced by dichlorvos (DDVP). We have previously shown organophosphate, DDVP-induced impairments in neurobehavioral indices viz. rota rod, passive avoidance, and water maze tests. In addition to this, we have also reported that chronic DDVP exposure leads to decreased mitochondrial electron transfer activities of cytochrome oxidase along with altered mitochondrial complexes I-III activity. Administration of CoQ(10) (4.5 mg/kg, i.p. for 12 weeks prior to DDVP administration daily) to DDVP-treated rats improved cognitive performance in passive avoidance task and Morris water maze test. Furthermore, CoQ(10) treatment also reduced oxidative stress (as evident by reduced malondialdehyde, decreased ROS and increased Mn-SOD activity) in DDVP-treated rats' hippocampus region, along with enhanced activity of complexes I-III and complex IV. Electron microscope studies of rat hippocampus mitochondria revealed that CoQ(10) administration leads to near normal physiology of mitochondria with well-defined cristae compared with DDVP-treated animals where enlarged mitochondria with distorted cristae are observed. CoQ(10) administration also attenuated neuronal damage in hippocampus as evident from histopathological studies. These results demonstrate the beneficial effects of CoQ(10) against organophosphate-induced cognitive impairments and hippocampal neuronal degeneration.

Apo-Eε4 allele in conjunction with Aß42 and tau in CSF: biomarker for Alzheimer's disease.

The objective of this study was to elucidate an association between Apo- Eε4 allele and CSF biomarkers Aß42 and tau for the diagnosis of Alzheimer's Disease (AD) patients. Aß42 and tau protein concentrations in CSF were measured by using ELISA assays. The levels of Aß42 were found to be decreased where as tau levels increased in AD patients. Moreover in AD patients Apo-Eε4 allele carriers have shown low Aß42 levels (328.86 ± 99.0 pg/ml) compared to Apo-Eε4 allele non-carriers (367.52 ± 5 7.37 pg/ml), while tau levels were higher in Apo-Eε4 allele carriers (511 ± 44.67 pg/ml) compared to Apo-Eε4 allele non-carriers (503.75 ± 41.08 pg/ml). Combination of Aß42 and tau resulted in sensitivity of 75.38% and specificity of 94.82% and diagnostic accuracy of 84.30% for AD compared with the controls. Therefore low Aß42 and elevated tau concentrations in CSF may prove to be a better diagnostic marker for AD along with the Apo-Eε4 allele.