Comparative Study of Accumulation of Two Potato Virus X Strains Differing in Virulence,Hydrolase Activity and Morphological Abnormalities in Datura stramonium L. Leaves

In young systemically infected leaves of Datura stramonium L., a severe strain of Potato virus X (PVX) accumulated to a lower degree than a mild strain. Infected leaves had increased protease and RNase activities in comparison with those of healthy controls. The highest hydrolase activities were found in leaves infected with the severe strain. Negative‐staining electron microscopy of dips from the infected leaves indicated that PVX virions underwent destructive changes, which resulted in the appearance of abnormal (swollen and ‘thin’) particles. Immuno‐electron microscopic assays showed that thin PVX particles, in contrast to those of normal diameter, lost the ability to bind with specific antiserum. The relative number of thin virions in leaves infected with the severe PVX strain was considerably higher than in leaves infected with the mild strain. This shows that a correlation exists between increased protease activity and intracellular destruction of virions. In abnormal virions, the viral RNA appears to be available for RNase attack. Therefore, it seems that high RNase activity together with increased generation of abnormal virions in the leaves infected with the severe strain promote inactivation of the viral RNA with RNase. We suppose that the enhanced hydrolase activities in the leaves infected with severe PVX strain, on the one hand, limit viral accumulation and thus play a defensive role and, on the other hand, cause considerable intracellular pathological changes resulting in severe symptoms.     

Apoptosis through Bcl-2/Bax and Cleaved Caspase Up-Regulation in Melanoma Treated by Boron Neutron Capture Therapy

Boron neutron capture therapy (BNCT) is a binary treatment involving selective accumulation of boron carriers in a tumor followed by irradiation with a thermal or epithermal neutron beam. The neutron capture reaction with a boron-10 nucleus yields high linear energy transfer (LET) particles, alpha and ⁷Li, with a range of 5 to 9 µm. These particles can only travel very short distances and release their damaging energy directly into the cells containing the boron compound. We aimed to evaluate proliferation, apoptosis and extracellular matrix (ECM) modifications of B16F10 melanoma and normal human melanocytes after BNCT. The amounts of soluble collagen and Hsp47, indicating collagen synthesis in the ECM, as well as the cellular markers of apoptosis, were investigated. BNCT decreased proliferation, altered the ECM by decreasing collagen synthesis and induced apoptosis by regulating Bcl-2/Bax in melanoma. Additionally, BNCT also increased the levels of TNF receptor and the cleaved caspases 3, 7, 8 and 9 in melanoma. These results suggest that multiple pathways related to cell death and cell cycle arrest are involved in the treatment of melanoma by BNCT.     

Identification and Prediction of Diabetic Sensorimotor Polyneuropathy Using Individual and Simple Combinations of Nerve Conduction Study Parameters

Objective

Evaluation of diabetic sensorimotor polyneuropathy (DSP) is hindered by the need for complex nerve conduction study (NCS) protocols and lack of predictive biomarkers. We aimed to determine the performance of single and simple combinations of NCS parameters for identification and future prediction of DSP.

Materials and Methods

406 participants (61 with type 1 diabetes and 345 with type 2 diabetes) with a broad spectrum of neuropathy, from none to severe, underwent NCS to determine presence or absence of DSP for cross-sectional (concurrent validity) analysis. The 109 participants without baseline DSP were re-evaluated for its future onset (predictive validity). Performance of NCS parameters was compared by area under the receiver operating characteristic curve (AROC).

Results

At baseline there were 246 (60%) Prevalent Cases. After 3.9 years mean follow-up, 25 (23%) of the 109 Prevalent Controls that were followed became Incident DSP Cases. Threshold values for peroneal conduction velocity and sural amplitude potential best identified Prevalent Cases (AROC 0.90 and 0.83, sensitivity 80 and 83%, specificity 89 and 72%, respectively). Baseline tibial F-wave latency, peroneal conduction velocity and the sum of three lower limb nerve conduction velocities (sural, peroneal, and tibial) best predicted 4-year incidence (AROC 0.79, 0.79, and 0.85; sensitivity 79, 70, and 81%; specificity 63, 74 and 77%, respectively).

Discussion

Individual NCS parameters or their simple combinations are valid measures for identification and future prediction of DSP. Further research into the predictive roles of tibial F-wave latencies, peroneal conduction velocity, and sum of conduction velocities as markers of incipient nerve injury is needed to risk-stratify individuals for clinical and research protocols.     

Effective gene flow in a historically fragmented area at the southern edge of silver fir (<Emphasis Type="Italic">Abies alba</Emphasis> Mill.) distribution

Fragmented populations at the edges of a species’ distribution can be highly exposed to the loss of genetic variation, unless sufficient gene flow maintains their genetic connectivity. Gene movements leading to successful establishment of external gametes (i.e. effective gene flow) into fragmented populations can solely be assessed by investigating the origin of natural regeneration. This study is focused on studying gene flow patterns in two silver fir stands in Central Apennines, where the species has a highly fragmented distribution. By using nuclear and chloroplast microsatellite markers, we investigated genetic variation, fine-scale spatial genetic structure, effective gene flow rates and large-scale connectivity characterizing both stands. Similar levels of genetic variation and low genetic differentiation between stands (F _ST = 0.005) and across generations were found, coupled with low inbreeding and weak to absent SGS in the adult cohort (Sp < 0.003). On the other hand, substantial differences between the two stands in terms of gene flow rates were observed. Irrespective of the parentage approach used, higher gene flow rates were found in the stand located at the upper silver fir altitudinal limit, especially for seed-mediated gene flow (0.79 in the upper stand vs. 0.48 in the lower stand). Conversely, the lower stand was characterized by a higher reproductive dominance of local adults. Our findings suggest that, despite similar levels of genetic variation and generally high gene flow rates, different processes may be acting on the two stands, reflecting varying ecological conditions.     

Reduction of apoptosis and preservation of mitochondrial integrity under ischemia/reperfusion injury is mediated by estrogen receptor β

Background

Estrogen improves cardiac recovery after ischemia/reperfusion (I/R) by yet incompletely understood mechanisms. Mitochondria play a crucial role in I/R injury through cytochrome c-dependent apoptosis activation. We tested the hypothesis that 17β-estradiol (E2) as well as a specific ERβ agonist improve cardiac recovery through estrogen receptor (ER)β-mediated mechanisms by reducing mitochondria-induced apoptosis and preserving mitochondrial integrity.

Methods

We randomized ovariectomized C57BL/6N mice 24h before I/R to pre-treatment with E2 or a specific ERβ agonist (ERβA). Isolated hearts were perfused for 20min prior to 30min global ischemia followed by 40min reperfusion.

Results

Compared with controls, ERβA and E2 treated groups showed a significant improvement in cardiac recovery, i.e. an increase in left ventricular developed pressure, dP/dtmax and dP/dtmin. ERβA and E2 pre-treatment led to a significant reduction in apoptosis with decreased cytochrome c release from the mitochondria and increased mitochondrial levels of anti-apoptotic Bcl2 and ACAA2. Protein levels of mitochondrial translocase inner membrane (TIM23) and mitochondrial complex I of respiratory chain were increased by ERβA and E2 pre-treatment. Furthermore, we found a significant increase of myosin light chain 2 (MLC2) phosphorylation together with ERK1/2 activation in E2, but not in ERβA treated groups.

Conclusions

Activation of ERβ is essential for the improvement of cardiac recovery after I/R through the inhibition of apoptosis and preservation of mitochondrial integrity and can be a achieved by a specific ERβ agonist. Furthermore, E2 modulates MLC2 activation after I/R independent of ERβ.