Biomarker System for Genetic Analysis of Stored Grain Pests Callosobruchus maculatus and Callosobruchus chinensis

Proceedings of the National Academy of Sciences, India Section B: Biological Sciences - Random amplified polymorphic DNA-PCR (RAPD-PCR) analysis was used in combination with morphological...     

Evaluation of in vitro effects of bevacizumab (Avastin) on retinal pigment epithelial, neurosensory retinal, and microvascular endothelial cells

PURPOSE: To evaluate the short-term in vitro safety of bevacizumab (Avastin) in human retinal pigment epithelial (ARPE-19), rat neurosensory retinal (R28), and human microvascular endothelial (HMVECad) cells. METHODS: ARPE-19 and R28 cells were treated with 0.125 mg/mL, 0.25 mg/mL, 0.50 mg/mL, and 1 mg/mL of bevacizumab for 2, 6, and 24 hours. HMVECad cells were treated with 5 ng/mL of vascular endothelial growth factor (VEGF) and 0.125 mg/mL, 0.25 mg/mL, 0.50 mg/mL, and 1 mg/mL of either bevacizumab for 2, 6, and 24 hours or a nonspecific human purified immunoglobulin (IgG) for 24 hours. Cell viability was measured using trypan blue dye exclusion assay. RESULTS: The cell viabilities of ARPE-19 cells, R28 cells, and HMVECad cells treated with bevacizumab were not significantly different (P > 0.05) from that of untreated controls. There was no significant difference (P > 0.05) between viabilities of HMVECad cells treated with bevacizumab and IgG. CONCLUSION: This study suggests that bevacizumab, at concentrations at or above the dose normally used in clinical practice, is not toxic to human retinal pigment epithelial, rat neurosensory retinal, or human microvascular endothelial cells in vitro. This report is consistent with the recent report of lack of toxicity of intravitreal bevacizumab in rabbits as well as the lack of apparent toxicity in clinical use.     

7-Ketocholesterol activates caspases-3/7, -8, and -12 in human microvascular endothelial cells in vitro

7-Ketocholesterol (7kCh) is a major oxysterol found associated with vascular diseases. Human microvascular endothelial cells (HMVECs) were cultured with different concentrations of 7kCh with and without inhibitors. Cell viabilities and caspase activities were assessed. 7kCh caused loss of cell viability in a dose-dependent manner. Caspases-8, -12, and -3/7 but not caspase-9 were activated by 7kCh treatment. The 7kCh-induced caspase-8 activity was blocked partially by pre-treatment with z-VAD-fmk and z-IETD-fmk, a caspase-8 inhibitor. However, pre-treatment with z-ATAD-fmk, a caspase-12 inhibitor, followed by 7kCh exposure lead to significantly increased caspase-8 activity. This suggests that caspase-8 and caspase-12 pathways have unique inhibition patterns and that caspase-12 is likely not upstream and feeding into caspase-8 but the pathways may function in parallel to each other. Caspase-3/7 activation was inhibited partially by low density lipoprotein (LDL), high density lipoprotein (HDL), z-VAD-fmk (pan-caspase inhibitor), and low doses (0.01 and 0.001 microM) of the cholesterol lowering drug, simvastatin. However, only LDL partially protected against 7kCh-induced loss of cell viability suggesting that caspase-independent pathways also contributed to the cell loss and that protection from oxysterol damage may require inhibition of multiple pathways. Moreover, our data suggest that oxysterols such as 7kCh can damage HMVECs cells in part via caspase-dependent apoptosis and may play a role in vascular and retinal diseases.     

Evaluation of levels of nitric oxide in saliva of children with rampant caries and early childhood caries: a comparative study

It is considered that caries incidence might be low in subjects with high salivary Nitric Oxide (NO) levels. Thus the objective of the present study was to determine the levels of nitric oxide in saliva of children with Rampant Caries (RC) and Early Childhood Caries (ECC). A total of 120 children were divided into 4 groups of 30 each belonging to two age groups of 6-12 yrs and 71 months or less respectively. Children between the age of 6-12 yrs were either with RC or their control and children between the age of 71 months or less were either with ECC or their control respectively. The study and control subjects were divided equally. Oral health status was recorded followed by unstimulated salivary flow rate estimation. Estimation of salivary nitric oxide was measured by the concentration of its stable metabolite nitrite using Classical Griess Reaction. The mean nitrite levels of both the control groups were much higher when compared with the study groups, which was statistically very highly significant.     

Application of second harmonic imaging microscopy to assess structural changes in optic nerve head structure ex vivo

Glaucoma represents the second leading cause of blindness worldwide. While both age and intraocular pressure (IOP) are well-recognized risk factors for this disease, the underlying pathologic process involves the accelerated death of retinal ganglion cells (RGCs) that is associated with progressive loss of vision. The loss of RGCs has been postulated to occur primarily by injury to axons in the optic nerve head (ONH) due to its anatomic features and the mechanical vulnerability of the lamina cribrosa, the specialized ONH zone comprised of collagen beams that define the channels or pores through which axon bundles exit the eye. Recent advances in multiphoton microscopy using femtosecond lasers that generate second harmonic (SH) signals from collagen allows for direct optical imaging of the lamina cribrosa. We assess the application of SH generated microscopy (SHG) to the study of the ONH, and test the general hypothesis that increasing intraocular pressure in the same eye results in the movement of ONH collagen beams leading to distortion of the lamina cribrosa channels and compression of the axon bundles.     

Caspase-8, -12, and -3 activation by 7-ketocholesterol in retinal neurosensory cells

PURPOSE: To determine the caspase pathways involved with 7-ketocholesterol (7kCh)-induced apoptosis in rat R28 cells. METHODS: R28 cells were exposed to 7kCh with or without low-density lipoprotein (LDL) and z-VAD-fmk, a pan-caspase inhibitor. Cell viability was measured by a trypan blue dye exclusion assay. Caspase-3, -8, -9, and -12 activities were measured by fluorochrome caspase assays. ARPE-19 cells were used as control for caspase-3 inhibition experiments. RESULTS: R28 cultures showed decreased cell viability on 7kCh exposure compared with controls (P < 0.001), and this was reversed with LDL and LDL + z-VAD-fmk (P < 0.001). The 7kCh-treated R28 cultures had increased caspase-8 activity compared with controls (P < 0.001). This activity was blocked partially with LDL (P < 0.01) or LDL + z-VAD-fmk (P < 0.001) but not with z-VAD-fmk alone. Caspase-12 activity was increased after 7kCh treatment compared with controls (P < 0.01), and this activity was increased further with the addition of LDL. Caspase-3 activity in R28 cultures increased with 7kCh treatment compared with controls (P < 0.001). In R28 cultures, the z-VAD-fmk treatment did not blocked 7kCh-induced caspase-3 activity but did block activity in ARPE-19 cultures (P < 0.001). Caspase-9 was not activated by 7kCh treatment. CONCLUSIONS: In R28 cells, 7kCh-induced apoptosis involves the caspase-3 along with the caspase-8 and caspase-12 pathways. LDL partially blocked 7kCh-induced caspase-8 activity but increased caspase-12 activities, suggesting that caspase-8 and caspase-12 pathways are independent of each other. The z-VAD-fmk inhibitor blocked caspase-3 activities in the homogeneous ARPE-19 cultures but not in the heterogeneous R28 cultures.