Association between the 4G/5G polymorphism of plasminogen activator inhibitor-1 (PAI-1) gene and sudden sensorineural hearing loss in Caucasian population: a meta-analysis.

PURPOSE: To clarify the association between the 4G/5G polymorphism of plasminogen activator inhibitor-1 (PAI-1) and sudden sensorineural hearing loss (SSNHL). METHODS: A systematic literature search of related studies up to August 30, 2019 in the PubMed and Embase databases was performed, and the results were displayed by odds ratios (ORs), and their 95% confidence intervals (CIs) were assessed using the STATA12.0 software using an allele model and a recessive model. RESULTS: Three eligible studies covering 519 subjects (241 cases, 278 controls) were identified. No statistically significant association was detected between the 4G/5G polymorphism and SSNHL in any model (allele model: 5G vs. 4G, OR = 0.952, 95% CI = 0.765-1.185, P = 0.662; recessive model: 5G/5G vs. 4G/5G + 4G/4G, OR = 0.841, 95% CI = 0.415-1.704, P = 0.631). CONCLUSIONS: There is no statistically significant association between the 4G/5G polymorphism of PAI-1 gene and SSNHL in the Caucasian population, and well-designed studies covering more patients and institutions should be conducted.

MiR-26a/miR-26b represses tongue squamous cell carcinoma progression by targeting PAK1.

BACKGROUND: Tongue squamous cell carcinoma (TSCC) is the most common oral malignancy. Previous studies found that microRNA (miR)-26a and miR-26b were downregulated in TSCC tissues. The current study was designed to explore the effects of miR-26a/miR-26b on TSCC progression and the potential mechanism. METHODS: Expression of miR-26a, miR-26b and p21 Activated Kinase 1 (PAK1) in TSCC tissues and cell lines was detected by reverse transcription- quantitative polymerase chain reaction (RT-qPCR). Flow cytometry analysis was performed to examine cell cycle and apoptosis. Transwell assay was conducted to evaluate the migrated and invasive abilities of SCC4 and Cal27 cells. In addition, western blot assay was employed to analyze the protein level. Glucose assay kit and lactate assay kit were utilized to analyze glycolysis. Dual-luciferase reporter and RNA immunoprecipitation (RIP) assays were applied to explore the relationship between miR-26a/miR-26b and PAK1. Xenograft tumor model was constructed to explore the role of miR-26a/miR-26b in vivo. RESULTS: Both miR-26a and miR-26b were underexpressed, while PAK1 was highly enriched in TSCC. Overexpression of miR-26a and miR-26b inhibited TSCC cell cycle, migration invasion and glycolysis, while promoted cell apoptosis. Both miR-26a and miR-26b directly targeted and negatively regulated PAK1 expression. Introduction of PAK1 partially reversed miR-26a/miR-26b upregulation-mediated cellular behaviors in TSCC cells. Gain of miR-26a/miR-26b blocked TSCC tumor growth in vivo. CONCLUSION: MiR-26a/miR-26b repressed TSCC progression via targeting PAK1 in vitro and in vivo, which enriched our understanding about TSCC development and provided new insights into the its treatment.

Substrate sustained release-based high efficacy biosynthesis of GABA by Lactobacillus brevis NCL912.

BACKGROUND: Gamma-aminobutyric acid (GABA) plays a significant role in the food and drug industries. Our previous study established an efficient fed-batch fermentation process for Lactobacillus brevis NCL912 production of GABA from monosodium L-glutamate; however, monosodium L-glutamate may not be an ideal substrate, as it can result in the rapid increase of pH due to decarboxylation. Thus, in this study, L-glutamic acid was proposed as a substrate. To evaluate its potential, key components of the fermentation medium affecting GABA synthesis were re-screened and re-optimized to enhance GABA production from L. brevis NCL912. RESULTS: The initial fermentation medium (pH 3.3) used for optimization was: 50 g/L glucose, 25 g/L yeast extract, 10 mg/L manganese sulfate (MnSO4·H2O), 2 g/L Tween-80, and 220 g/L L-glutamic acid. Glucose, a nitrogen source, magnesium, and Tween-80 had notable effects on GABA production from the L-glutamic acid-based process; other factors showed no or marginal effects. The optimized levels of the four key components in the fermentation medium were 25 g/L glucose, 25 g/L yeast extract FM408, 25 mg/L MnSO4·H2O, and 2 g/L Tween-80. A simple and efficient fermentation process for the bioconversion of GABA by L. brevis NCL912 was subsequently developed in a 10 L fermenter as follows: fermentation medium, 5 L; glutamic acid, 295 g/L; inoculum, 10% (v/v); incubation temperature, 32 °C; and agitation, 100 rpm. After 48 h of fermentation, the final GABA concentration increased up to 205.8 ± 8.0 g/L. CONCLUSIONS: L-Glutamic acid was superior to monosodium L-glutamate as a substrate in the bioproduction of GABA. Thus, a high efficacy bioprocess with 205 g/L GABA for L. brevis NCL912 was established. This strategy may provide an alternative for increasing the bioconversion of GABA.

miR-375-3p inhibits the progression of laryngeal squamous cell carcinoma by targeting hepatocyte nuclear factor-1ß.

Laryngeal squamous cell carcinoma (LSCC) is one of the most frequently diagnosed head and neck cancers worldwide. Increasing evidence suggests that microRNAs (miRNAs/miRs) regulate the progression of tumorigenesis and the malignant behaviors of cancer cells. The aim of this study was to investigate the function and underlying mechanism of miR-375-3p in LSCC. The expression of miR-375-3p in LSCC tissues and cells was detected using reverse transcription-quantitative PCR. The effects of miR-375-3p on the malignant phenotype of LSCC cells was determined using the Cell Counting Kit-8 assay and flow cytometry. The targets of miR-375-3p were predicted using the miRDB database and confirmed by the luciferase reporter assay. The results of the present study demonstrated that miR-375-3p was downregulated in LSCC tissues and cell lines. Furthermore, overexpression of miR-375-3p significantly suppressed the proliferation and cell cycle progression of LSCC cells. Overexpression of miR-375-3p also increased LSCC cell apoptosis. Mechanistical analysis indicated that miR-375-3p bound the 3'-untranslated region of the hepatocyte nuclear factor 1ß (HNF1ß) and decreased its expression in LSCC cells. Consistent with the role of HNF1ß in glucose metabolism, overexpression of miR-375-3p significantly inhibited glucose consumption and lactate production in LSCC cells. Transfection with HNF1ß notably reversed the inhibitory effect of miR-375-3p on the proliferation of LSCC cells. Collectively, these results indicate the tumor suppressive role of miR-375-3p in LSCC via HNF1ß, suggesting that miR-375-3p may serve as a potential target in the treatment of LSCC.

Hsa_circ_0042666 inhibits proliferation and invasion via regulating miR-223/TGFBR3 axis in laryngeal squamous cell carcinoma.

Circular RNAs (circRNAs) have been reported to play critical roles in tumorigenesis. However, the roles of circRNAs in laryngeal squamous cell carcinoma (LSCC) are still largely unknown. In our present study, we identified a novel circRNA hsa_circ_0042666, which was poorly expressed in LSCC. Low hsa_circ_0042666 expression was closely associated with advanced tumor stage, lymph-node metastasis, and poor overall survival. In vitro function assays, we showed that hsa_circ_0042666 dramatically reduced LSCC cells proliferation and invasion in vitro. In mechanism, our data indicated that hsa_circ_0042666 could competitively bind to miR-223 as a miRNA sponge to regulate TGFBR3 expression in LSCC progression. Altogether, these findings elucidated that hsa_circ_0042666 regulated LSCC cells proliferation and invasion by miR-223/TGFBR3 axis, which might provide a therapeutic strategy for the treatment of LSCC.

Disassociation of glutamate from γ-aminobutyric acid by zinc acetate-assisted differential precipitation/dissolution: Application to the quantification of γ-aminobutyric acid.

γ-aminobutyric acid (GABA) is a key physiologically active molecule in organisms. Separation of glutamate from its decarboxylated product GABA has been vigorously pursued. The interaction between these two compounds severely hindered their disassociation. Herein, we present a new strategy, termed zinc acetate-assisted differential precipitation/dissolution (ZA-DPD), for the removal of glutamate by step by step recovering pure GABA solution and discarding pure glutamate pellet, essentially attributed to the use of two core reagents (zinc acetate-assisted glutamate-precipitating reagent, and glutamate-rejecting reagent). In each precipitation, the zinc acetate-assisted glutamate-precipitating reagent guaranteed most GABA still soluble although the rest co-precipitated with glutamate; in the coupled dissolution, the co-precipitated GABA was fully dissolved with or without (in the case of glutamate-rejecting reagent used in the final dissolution) co-dissolution of glutamate. The process was repeated twice until glutamate was thoroughly removed. An accurate quantitative method coupling ZA-DPD with colorimetry was thereafter established for the determination of GABA. This study may facilitate the areas associated with GABA or glutamate.

Genomic insights into a robust gamma-aminobutyric acid-producer Lactobacillus brevis CD0817.

Lactobacillus brevis CD0817, a strain isolated from a healthy adult gut, was currently the most efficient lactic acid bacterial cell factory for gamma-aminobutyric acid. In this study, the complete genome sequence of CD0817 was determined and compared with some related L. brevis genomes. The CD0817 genome consists of one 2,990,570-bp chromosome and four plasmids. The comparative genomic and phylogenetic analysis revealed that L. brevis CD0817 was not very conserved with low GABA-producing L. brevis strains. A significant divergence was that CD0817 harbors only the gadCA operon whereas the low GABA-producing L. brevis strains contain the operon and gadB. The gadB seemed to only marginally contribute to the accumulation of GABA. The high GABA production ability of CD0817 may be associated with its extraordinary genome.

Stepwise partially overlapping primer-based PCR for genome walking.

A stepwise partially overlapping primer-based PCR (SWPOP-PCR) method for isolating flanking unknown DNA regions was developed, which comprises three rounds of nested PCRs sequentially driven by SWPOP primer-nested specific primer pairs. SWPOP primer set is characterized by a partial overlap of 10 bp with 3'-part of the latter primer is identical to 5'-part of the former one, which makes the SWPOP primer in use anneal to SWPOP site of the prior PCR product only at relatively low temperature. For each PCR, target single-stranded DNA primed by the SWPOP primer in the exclusive one low-stringency cycle is converted into double-stranded form in the following high-stringency cycle due to the presence of a perfect annealing site for the specific primer. This double-stranded DNA bounded by the specific primer and the SWPOP primer is exponentially amplified in the remaining high-stringency cycles. Non-target single-stranded DNA, however, cannot be amplified given the lack of perfect complementary sequences for any primers. Therefore, the partial overlap of a SWPOP primer set preferentially synthesizes target products but inhibits nonspecific amplification. We successfully exploited SWPOP-PCR to obtain the DNA sequences flanking glutamate decarboxylase gene (gadA) locus in Lactobacillus brevis NCL912 and hygromycin gene (hyg) integrated in rice.