Downregulation of lncRNA APCDD1L-AS1 due to DNA hypermethylation and loss of VHL protein expression promotes the progression of clear cell renal cell carcinoma

Background: The current studies only indicated that long non-coding RNA (lncRNA) APCDD1L-AS1, as a novel lncRNA, may play a role in oral squamous cell carcinoma and lung cancer. However, its potential role in clear cell renal cell carcinoma (ccRCC) and its possible mechanism of action remain vague.Methods: TCGA-KIRC and GEO data and qRT-PCR and pyrosequencing results of clinical specimens were used to identify the expression level and DNA methylation status of APCDD1L-AS1. The effects of APCDD1L-AS1 overexpression on ccRCC growth and metastasis were determined by function experiments. Western blot and Tandem mass tags (TMT) were utilized to explore the relationship between APCDD1L-AS1 and VHL expression and its downstream underlying mechanisms.Results: The expression of APCDD1L-AS1 was downregulated in ccRCC. Decreased APCDD1L-AS1 expression was related to higher tumor stage and histological grade and shorter RFS (Relapse-free survival). Besides, APCDD1L-AS1 overexpression restrained the growth and metastasis of ccRCC cells in vitro and in vivo. Moreover, reduced APCDD1L-AS1 expression could be caused by DNA hypermethylation and loss of von Hippel Lindau (VHL) protein expression. Furthermore, the dysregulation of histones expression caused by APCDD1L-AS1 overexpression may be one of the important mechanisms to suppress the progression of ccRCC.Conclusion: APCDD1L-AS1 was able to inhibit the progression of ccRCC, and its decreased expression could be caused by DNA hypermethylation and loss of VHL protein expression. Therefore, APCDD1L-AS1 may serve as a new therapeutic target in the treatment of ccRCC.     

Investigation of arcobacters in meat and faecal samples of clinically healthy cattle in Turkey

AIMS: To investigate the presence of Arcobacter spp. in minced beef meat (n = 97) and rectal faecal samples (n = 200) collected from cattle immediately after slaughter at a local abattoir in Turkey. METHODS AND RESULTS: Meat samples were examined using three different isolation procedures (CAT-supplemented media, de Boer arcobacter isolation method and membrane filtration method), but only one method (CAT-supplemented media) was employed for faecal samples. The isolated Arcobacter strains were identified by genus- and species-(multiplex) specific PCR assays. Arcobacter spp. were isolated from 5 and 9.5% of meat and faecal samples respectively. Although the only Arcobacter sp. found in meat samples was Arcobacter butzleri, all three pathogenic species--A. butzleri, A. cryaerophilus and A. skirrowii--were detected in the rectal swabs. No Arcobacter was isolated when the de Boer method was used for minced meat samples but the same five meat samples were found positive for arcobacters when CAT-supplemented media and membrane filtration method were used. CONCLUSIONS: The membrane filtration method was found to be superior to the CAT-supplemented media, because it led to a reduction in competing microflora. However, the necessity for one filter and medium for each sample makes this method somewhat expensive. The multiplex-PCR (m-PCR) assay shortened significantly the time required for the identification of Arcobacter spp. and also removed the possibility of false positive results due to other campylobacteria. SIGNIFICANCE AND IMPACT OF THE STUDY: This study reports the isolation of Arcobacter spp. in cattle for the first time in Turkey. The m-PCR assay enables the identification and differentiation of all arcobacters simultaneously in one-step PCR.     

Validity of two alternative systems for measuring vertical jump height

Vertical jump height is frequently used by coaches, health care professionals, and strength and conditioning professionals to objectively measure function. The purpose of this study is to determine the concurrent validity of the jump and reach method (Vertec) and the contact mat method (Just Jump) in assessing vertical jump height when compared with the criterion reference 3-camera motion analysis system. Thirty-nine college students, 25 females and 14 males between the ages of 18 and 25 (mean age 20.65 years), were instructed to perform the countermovement jump. Reflective markers were placed at the base of the individual's sacrum for the 3-camera motion analysis system to measure vertical jump height. The subject was then instructed to stand on the Just Jump mat beneath the Vertec and perform the jump. Measurements were recorded from each of the 3 systems simultaneously for each jump. The Pearson r statistic between the video and the jump and reach (Vertec) was 0.906. The Pearson r between the video and contact mat (Just Jump) was 0.967. Both correlations were significant at the 0.01 level. Analysis of variance showed a significant difference among the 3 means F(2,235) = 5.51, p < 0.05. The post hoc analysis showed a significant difference between the criterion reference (M = 0.4369 m) and the Vertec (M = 0.3937 m, p = 0.005) but not between the criterion reference and the Just Jump system (M = 0.4420 m, p = 0.972). The Just Jump method of measuring vertical jump height is a valid measure when compared with the 3-camera system. The Vertec was found to have a high correlation with the criterion reference, but the mean differed significantly. This study indicates that a higher degree of confidence is warranted when comparing Just Jump results with a 3-camera system study.     

Pin1 inhibition activates cyclin D and produces neurodegenerative pathology

Abnormal cell cycle events are increasingly becoming important attributes of neurodegenerative pathology. Pin1 is a crucial target of neurodegeneration in relation to its functions regarding these abnormal cell cycle events in neurons. Pin1 is majorly involved in many aspects of cell cycle regulation and it has also been suggested to have a neuroprotective function against neurodegenerative pathologies. Oxidative dysregulation of Pin1 affects not only normal tau regulation, eventually causing tangle formation, but also cell cycle regulation in neurons. Presence of cell cycle proteins has been shown in many neurodegenerative diseases. Importantly, many of these proteins have physical interactions with Pin1. Hence, understanding Pin1's role in abnormal cell cycle re-entry is critical in terms of finding new approaches for the future therapeutic options treating neurodegenerative pathologies. Here, we show that inhibition of Pin1 by its selective inhibitor juglone leads to up-regulation of cyclinD1, phospho-tau, and caspase 3, producing apoptosis in cultured rat hippocampal neurons. We also observed axonal retraction with a change in sub-cellular localizations of cyclins. Therefore, Pin1 dysregulation, in relation to its role in cell cycle regulation in neurons, may have profound effects in the progression of neurodegenerative pathology, making it a possible crucial target behind many neurodegenerative diseases.     

Differentiation of Arcobacter species by numerical analysis of AFLP profiles and description of a novel Arcobacter from pig abortions and turkey faeces

AIMS: To evaluate the efficacy of amplified fragment length polymorphism (AFLP)-based genetic profiling for taxonomic and epidemiological analyses of diverse Arcobacter species. METHODS AND RESULTS: Seventy-two isolates of A. butzleri, A. cryaerophilus, A. skirrowii and A. nitrofigilis, and a previously unclassified porcine abortion strain were studied. AFLP profiling was performed using a BglII-Csp6I-based protocol previously used to characterize Campylobacter species. Duplicate profiles of 20 isolates were 93.25% similar, indicating high reproducibility. Numerical analysis of all 72 strains revealed five phenons at the 29% similarity level, four of which represented each of the known species studied. The remaining phenon was further characterized by phenotypic and 16S rDNA sequence analyses, the results of which indicated it to be a novel Arcobacter species. The genetically distinct subgroups of A. cryaerophilus were differentiated at the 39.5% similarity level. For strain typing, 62 distinct types were defined, with evidence of clonal lineages within A. butzleri, A. cryaerophilus and A. skirrowii. CONCLUSIONS: AFLP profiling is an effective means of determining taxonomic and strain relationships for arcobacters. SIGNIFICANCE AND IMPACT OF THE STUDY: First use of AFLP profiling for diverse Arcobacter species; indication of clonality in A. butzleri, A. cryaerophilus and A. skirrowii; potentially novel Arcobacter taxon identified.     

The lambda Gam protein inhibits RecBCD binding to dsDNA ends

Inactivation of the Escherichia coli RecBCD enzyme by the lambda Gam protein is an essential step that accompanies the lambda Red proteins for gene replacement using recombineering technology. It has been shown that Gam inhibits all the activities of RecBCD to the same extent. Nonetheless, some in vivo properties of recBCD mutants cannot be mimicked effectively by the expression of gam in vivo. An examination of the mechanism of Gam's inhibition of RecBCD was performed, and it was found that Gam inhibits the binding of RecBCD to double-stranded DNA ends, even if RecBCD is bound to DNA before its interaction with Gam. When ATP is added to the reaction to induce helicase activity, most of the reaction is inhibited by Gam, but residual amounts of unwinding are detected, despite a 40-fold excess of Gam/RecBCD. The same inhibitory effect of Gam was seen on RecBCD that had been modified by the P22 anti-RecBCD protein Abc2, though the inhibitory effect was diminished due to the tighter binding of Abc2-modified RecBCD to double-stranded DNA ends. These data suggest that cells containing Gam-expressing plasmids retain a small amount of uninhibited enzyme. Given the suspected instability of Gam in vivo, care must be taken when interpreting results from experiments containing Gam-inhibited RecBCD species. A revised model is proposed for Gam-induced radioresistance of E. coli to ionizing radiation.     

Cyclic AMP is sufficient for triggering the exocytic recruitment of aquaporin-2 in renal epithelial cells

The initial response of renal epithelial cells to the antidiuretic hormone arginine vasopressin (AVP) is an increase in cyclic AMP. By applying immunofluorescence, cell membrane capacitance and transepithelial water flux measurements we show that cAMP alone is sufficient to elicit the antidiuretic cellular response in primary cultured epithelial cells from renal inner medulla, namely the transport of aquaporin-2 (AQP2)-bearing vesicles to, and their subsequent fusion with, the plasma membrane (AQP2 shuttle). The AQP2 shuttle is evoked neither by AVP-independent Ca²⁺ increases nor by AVP-induced Ca²⁺ increases. However, clamping cytosolic Ca²⁺ concentrations below resting levels at 25 nM inhibited exocytosis. Exocytosis was confined to a slow monophasic response, and readily releasable vesicles were missing. Analysis of endocytic capacitance steps revealed that cAMP does not decelerate the retrieval of AQP2 from the plasma membrane. Our data suggest that cAMP initiates an early step, namely the transport of AQP2-bearing vesicles towards the plasma membrane, and do not support a regulatory function for Ca²⁺ in the AQP2 shuttle.