Preparation and Initial Application of a Monoclonal Antibody Specific for a Newly Discovered Conserved Linear Epitope of Rabies Virus Nucleoprotein

Objective To prepare monoclonal antibodies against a newly discovered and conserved linear epitope of Rabies virus nucleoprotein and to use them in a rabies diagnostic test. Methods Synthetic peptide containing the epitope was used as immunogen to prepare hybridoma cell lines by classical hybridoma technology. Anti-peptide monoclonal antibodies produced in ascites of inoculated Balb/c mice were labeled with fluorescein isothiocyanate （FITC） after purification and used in fluorescent antibody test （FAT）. Results Two positive hybridoma cell lines, RVNP-mAbl-CL and RVNP-mAb2-CL, were obtained. RVNP- mAbl-CL produced a higher concentration of monoclonal antibody RVNP-mAbl in Balb/c ascites. FITC-labeled RVNP-mAbl showed correct results on certain Rabies virus-positive canine brain tissue samples and cells of a small subclone of baby hamster kidney 21 cell line （BSR）. Conclusion FITC-labeled RVNP-mAbl has potential application for laboratory diagnosis of rabies

Development of an Internally Controlled Reverse Transcription Recombinase-aided Amplification Assay for the Rapid and Visual Detection of West Nile Virus

West Nile virus(WNV)causes West Nile fever and West Nile encephalitis.Because infection by WNV creates serious public health problems,its simple,rapid,and visual detection is very important in clinical practice,especially in resource-limited laboratories.We have developed a rapid,specific,and highly sensitive internally controlled reverse transcription recombinase-aided amplification(RTRAA)assay to detect WNV,using both real-timefluoresce nee and the lateral flow dipstick(LFD)at39.0°C for 30 min.The analytical sensitivity of theRT-RAA assay was 10 plasmid copies and 1.6 pfu perreacti on with real-time fluoresce nee,and 1,000plasmid copies per reaction with the LFD.No crossreactionwith other control viruses was observed.Compared with the RT-qPCR assay,the RT-RAA assaydemonstrated 100%sensitivity and 100%specificityfor WNV.

Yes-associated protein at the intersection of liver cell fate determination

A recent publication highlights the importance of high yes-associated protein(YAP) expressing cells in liver regeneration following partial hepatectomy.Although the names of the cell populations described in these articles [hybrid periportal hepatocytes(HybHP) or epithelial-mesenchymal transition(EMT)-reprogrammed hepatocytes] are not identical, they all express high levels of YAP.We hypothesize that the HybHP and EMT-reprogrammed hepatocytes might be a similar cell population. Hippo signaling is the primary pathway that regulates YAP activity. According to the contribution of these two types of cells to liver regeneration and the high YAP expression, Hippo-YAP signaling activation may be a common regulatory pathway experienced by cells undergoing dedifferentiation and reactivating proliferative activity during liver regeneration.Although no evidence has shown that HybHP cells contribute to hepatocellular carcinoma in mouse models, we can not rule out the possibility that these highly regenerative cells can further develop into tumor cells when they acquire mutations caused by viral infection or other risk factors like alcohol. The detailed mechanistic insight of the regulation of YAP expression and activity in HybHP(or other types of cells contributing to liver regeneration) is unknown. We hypothesize that liver regeneration under various conditions will eventually lead to divergent consequences, likely due to the duration of YAP activation regulated by Hippo-large tumor suppressor 1 and 2 pathway in a context-and cell typedependent manner.

Toll-like receptor 10 (TLR10) exhibits suppressive effects on inflammation of prostate epithelial cells

Prostate inflammation (PI) is closely related to the development and progression of chronic prostatic diseases: benign prostatic hyperplasia and prostate cancer. Toll-like receptor (TLR) 2 has been reported to be associated with inflammatory diseases, such as infections, autoimmune diseases, and cancers. Meanwhile, TLR10, which can form heterodimers with TLR2, has been considered an orphan receptor without an exact function. The present study therefore aims to examine the effects of TLR2 and TLR10 on PI. Prostate samples and clinical data were obtained from the patients diagnosed with benign prostatic hyperplasia. The inflammatory cell model was established by adding lipopolysaccharide to RWPE-1 cells. Prostate tissues/cells were examined by histological, molecular, and biochemical approaches. Both TLR2 and TLR10 were found to be expressed in prostate tissues and RWPE-1 cells. mRNA/protein expression levels of TLR2 and TLR10 were both positively correlated with prostate tissue inflammatory grades. Lipopolysaccharide-stimulated RWPE-1 cells expressed higher levels of TLR2, TLR10, high mobility group box 1 (HMGB1), phosphonuclear factor kappa-light-chain-enhancer of activated B-cells P65 (phospho-NF-κB P65), interleukin (IL)-6, and IL-8 than control cells. Moreover, HMGB1, phospho-NF-κB P65, IL-6, and IL-8 were down regulated after TLR2 knockdown and upregulated after TLR10 knockdown in RWPE-1 cells. TLR2 stimulation can activate the inflammatory signaling cascade in prostate epithelial cells. Conversely, TLR10 exhibited suppressive effects on inflammation. With antagonistic functions, both TLR2 and TLR10 were invoIved in PI. TLR10 could be a novel target in modulating inflammatory signal transduction of prostate epithelial cells.

Active Compounds and Molecular Targets of Chinese Herbal Medicine for Neurogenesis in Stroke Treatment: Implication for Cross Talk between Traditional Chinese Medicine and Biomedical Sciences

Neural stem/progenitor cells (NSCs) could be attractive therapeutic targets for promoting adult neurogenesis, brain plasticity, and repair in stroke and neurodegenerative diseases, raising great potentials for regeneration therapy. In adult ischemic brains, NSCs have limited capacities of growth, differentiation, and generating new neurons for repairing the damaged central nervous system. However, the spontaneous brain repair seems to be insufficient to recover neurological deficits in most stroke cases. To overcome those problems, pharmacological manipulations targeting on endogenous NSCs or transplanted stem cells could be a promoting strategy for regeneration therapy. Chinese herbal medicine has great potentials for developing novel therapeutic strategies for adult neurogenesis and brain repair in poststroke treatment. Chinese herbal medicine has a long history for poststroke treatment. Recent studies create great opportunity for drug discovery for promoting neurogenesis and improving the recovery of neurological fimctions in poststroke treatment. Many active compounds or extracts from medicinal herbs have shown promising effects on regulating proliferation, self-renewal and differentiation of NSCs, and promoting neural network formation as well as neurological functional recovery with in vitro and in vivo experimental evidence. Therefore, targeting neural stem/progenitor cells can be an important opportunity for the studies of Traditional Chinese Medicine in regeneration medicine. Due to the complex interactions of herbal ingredients in network regulation, huge challenge remains to be resolved for further study.

Diagnosis of renal amyloidosis by liquid chromatography-tandem mass spectrometry:experience from a single-center cohort study in China

To the Editor:Amyloidosis is caused by the extracellular deposition of insoluble beta-pleated protein fibrils in tissues and is classified according to the constituent proteins.The cause of amyloidosis can be traced to more than 30 kinds of malfolded proteins,and accurate amyloid typing is crucial for appropriate therapy.[1]The pathological diagnosis is confirmed with positive Congo red(CR)staining under a light microscope and 7 to 12 nm branching fibrils under an electron microscope.Further identification of some amyloid precursor proteins can resort to immunofluorescence(IF)and immunochemistry(IHC);however,the accuracy depends on the selected antibodies to a large extent.Laser microdissection coupled with liquid chromatography-tandem mass spectrometry(LMD/MS)can identify these proteins without the restriction of IF and IHC,although exquisite equipment and skilled specialists are required.[2]This study aimed to establish LMD/MS analysis of renal biopsy tissue and compare it with IHC on diagnostic sensitivity.

Ficolin A exacerbates severe H1N1 influenza virus infection-induced acute lung immunopathological injury via excessive complement activation

Severe pandemic influenza A(H1N1)2009 virus(pH1N1)infection causes significant morbidity and mortality.We and other groups have reported that immunopathological damage induced by excessive pulmonary inflammation plays a critical role in the pathogenesis of severe pneumonia and provides novel strategies for the treatment of severe influenza infection[1–3].The complement system plays critical roles in both innate and adaptive immunity by activating the classical,alternative and lectin pathways[4].