The efficacy and safety of fexuprazan in treating erosive esophagitis: a phase III, randomized, double-blind, multicenter study.

BACKGROUND AND AIM: Fexuprazan is a novel potassium-competitive acid blocker (P-CAB). This study aimed to explore the noninferior efficacy and safety of fexuprazan to esomeprazole in treating erosive esophagitis (EE). METHODS: This was a phase III, randomized, double-blind multicenter study. Patients with endoscopically confirmed EE were randomized to receive fexuprazan 40 mg or esomeprazole 40 mg once a daily for 4-8 weeks. The healing rates of EE, symptom response, GERD-health-related quality life (GERD-HRQL), and treatment-emergent adverse events (TEAEs) were compared between fexuprazan group and esomeprazole group. RESULTS: A total of 332 subjects were included in full analysis set (FAS) and 311 in per-protocol set (PPS). The healing rates of fexuprazan and esomeprazole groups at 8 weeks were 88.5% (146/165) and 89.0% (145/163), respectively, in FAS and 97.3% (145/149) and 97.9% (143/146), respectively, in PPS. Noninferiority of fexuprazan compared with esomeprazole according to EE healing rates at 8 weeks was demonstrated in both FAS and PPS analysis. No significant difference was found between groups in EE healing rates at 4 weeks, symptom responses, and changes of GERD-HRQL. The incidence of drug-related AEs was 19.4% (32/165) in fexuprazan arm and 19.6% (32/163) in esomeprazole arm. CONCLUSION: This study demonstrated noninferior efficacy of fexuprazan to esomeprazole in treating EE. The incidence of TEAEs was similar between fexuprazan and esomeprazole. Trial registration number NCT05813561.

Genome-wide identification and characterization of superoxide dismutases in four oyster species reveals functional differentiation in response to biotic and abiotic stress.

BACKGROUND: Oysters inhabit in the intertidal zone and may be suffered from environmental stresses, which can increase the production of reactive oxygen species (ROS), resulting in mass mortality. Superoxide dismutases (SODs) protect oysters from ROS damage through different mechanisms compared with vertebrates. However, the molecular and functional differentiation in oyster SODs were rarely analyzed. RESULT: In this study, a total of 13, 13, 10, and 8 candidate SODs were identified in the genome of Crassostrea gigas, Crassostrea virginica, Crassostrea hongkongensis, and Saccostrea glomerata respectively. The domain composition, gene structure, subcellular locations, conserved ligands, and cis-elements elucidated the SODs into five groups (Mn-SODs, Cu-only-SODs, Cu/Zn ion ligand Cu/Zn-SOD with enzyme activity, Zn-only-SODs, and no ligand metal ions Cu/Zn-SODs). For single domain Cu/Zn-SODs, only one cytosolic Cu/Zn-SOD (cg_XM_034479061.1) may conserve enzymatic activity while most extracellular Cu/Zn-SOD proteins appeared to lose SOD enzyme activity according to conserved ligand amino acid analysis and expression pattern under biotic and abiotic stress in C. gigas. Further, multi-domain-SODs were identified and some of them were expressed in response to biotic and abiotic stressors in C. gigas. Moreover, the expression patterns of these genes varied in response to different stressors, which may be due to the cis-elements in the gene promoter. CONCLUSION: These findings revealed the most extracellular Cu/Zn-SOD proteins appeared to lose SOD enzyme activity in oysters. Further, our study revealed that only one cytosolic Cu/Zn-SOD (cg_XM_034479061.1) may conserve enzymatic activity of SOD. Moreover, the expression patterns of these genes varied in response to different stressors, which may be due to the cis-elements in the promoter. This study provides important insights into the mechanisms through which oysters adapt to harsh intertidal conditions, as well as potential biomarkers of stress response in related species.

Transcriptomic identification of key genes in Pacific oysters Crassostrea gigas responding to major abiotic and biotic stressors.

Oysters are commercially important intertidal filter-feeding species. Mass mortality events of oysters often occur due to environmental stresses, such as exposure to fluctuating temperatures, salinity, and air, as well as to metal pollution and pathogen infection. Here, RNA-seq data were used to identify shared and specific responsive genes by differential gene expression analysis and weighted gene co-expression network analysis. A total of 18 up-regulated and 10 down-regulated shared responsive genes were identified corresponding to five different stressors. Total 27 stressor-specific genes for temperature, 11 for salinity, 80 for air exposure, 51 for metal pollution, and 636 for Vibrio mediterranei pathogen stress were identified in oysters. Elongin-ß was identified as a crucial gene for thermal stress response. Some HSP70s were determined to be shared responsive genes while others were specific to thermal tolerance. The proteins encoded by these stress-related genes should be further investigated to characterize their physiological functions. In addition, the uncharacterized proteins and ncRNAs that were identified may be involved in species-specific stress-response and regulatory mechanisms. This study identified specific genes related to stressors relevant to oyster cultivation. These findings provide useful information for new selective breeding strategies using a data driven method.

The complete mitochondrial genome of Crassostrea hongkongensis from East China Sea indicates species' range may extend northward.

BACKGROUND: Crassostrea hongkongensis is an important mariculture shellfish with a relatively narrow distribution range. Recently, larger wild oysters were identified as C. hongkongensis from Sanmen bay in East China Sea. No natural distribution had been reported for this species here, and its origin remains unknown. METHODS AND RESULTS: We assembled the complete 18,617 bp circular mitochondrial genome of C. hongkongensis from Sanmen bay by next generation sequencing. It included 12 protein-coding genes, 23 tRNAs, and two rRNAs. The A/T content of the mitogenome was higher than its G/C content. Similar values and features were previously found for five other specimens of C. hongkongensis, and were comparable to those of other congeneric species. A phylogenetic analysis based on the 12 protein-coding genes and complete mitochondrial sequence indicated that the six specimens of C. hongkongensis formed a monophyletic group and shared a sister group relationship with C. ariakensis, C. nippona, C. sikamea, C. angulata, C. gigas, and C. iredalei, whereas specimens from the Sanmen bay area clustered later with the five other C. hongkongensis individuals, sharing a sub-clade. The newly sequenced mitogenome had more singleton sites than previously published C. hongkongensis mitogenomes. CONCLUSIONS: Crassostrea hongkongensis may be a native species, and the species' range extends further to the north than previously known. Our data may therefore contribute to a better understanding of the species diversity and conservation of Crassostrea oysters.

RNAi based transcriptome suggests genes potentially regulated by HSF1 in the Pacific oyster Crassostrea gigas under thermal stress.

BACKGROUND: The Pacific oyster Crassostrea gigas is an important fishery resource that is sensitive to temperature fluctuations. Thus, it has evolved a protection mechanism against heat stress by increasing the expression of the gene coding for heat shock protein (HSP) 70 under elevated temperatures. In other animals, heat shock response is a transcriptional response driven by the heat shock transcription factor 1 (HSF1) and thermal stress can trigger HSP70 expression to protect the organism via HSF1. However, the regulatory relationship between HSF1 and HSP remains unclear in Pacific oyster. Therefore, in the present study, we examined the transcriptomic response of several to thermal stress following HSF1 interference. RESULTS: We identified 150 genes responsive to heat shock including seven HSP genes, six of which belonging to the group of 17 HSP genes enriched in response to heat shock, according to weighted gene co-expression network analysis (WGCNA). The other gene was enriched in the module correlated with HSF1 interference. In addition, we found 48 and 47 genes that were upregulated and downregulated by HSF1 in response to heat shock, respectively. In the upregulated genes, we identified one HSP70 potentially regulated by HSF1 in response to heat shock. Furthermore, based on differentially expressed genes and WGCNA analyses, we found that the hypoxia signaling pathway was enriched under heat shock conditions. Five genes were then selected to detect dynamic changes through time. The results suggested that gene expression was correlated with HSF1 expression. The regulation of HSP70 by HSF1 was preliminarily confirmed by binding site predictions and by a dual luciferase assay. CONCLUSIONS: Our results revealed that the expression of HSP70 and HSP20 was initially triggered after 2 h of heat shock, and one of the HSP70 genes was potentially regulated by HSF1. From these results, it is evident that not all heat-inducible genes were triggered simultaneously in response to heat shock stress. Overall, the results revealed a possible HSF1-HSP regulatory relationship in Pacific oyster, providing valuable information on the mechanisms of thermal tolerance in this commercially important oyster.

[Significance of the items for Dizziness Handicap Inventory in differential diagnosis of benign paroxysmal positional vertigo].

OBJECTIVE: To explore the value of items for the Chinese version of Dizziness Handicap Inventory (DHI) in differential diagnosis of benign paroxysmal positional vertigo (BPPV) in patients with vertigo or dizziness first coming to the outpatient clinic.
 Methods: A total of 322 patients with vertigo or dizziness, who came from Nanfang Hospital, Southern Medical University, were enrolled from April 2016 to February 2017. The Chinese version of DHI and Visual Analogue Scale (VAS) were completed by themselves. After detailed vestibular function examination, the patients were divided into a BPPV group, a normal vestibular group, and a abnormal vestibular group.
 Results: The score of DHI-2 in the BPPV group was 5.52±2.10, which was higher than that in the normal vestibular group (3.94±2.91)(t=3.847, P<0.01) and the abnormal vestibular group (4.17±2.74)(t=4.149, P<0.01). There were significant differences in the DHI-2 among the 3 groups (F=9.870, t=4.515, P<0.01). The score of DHI-item 13 in the BPPV group was 3.09±1.39, which was higher than that in the normal vestibular group (1.97±1.63)(t=4.515, P<0.01) and the abnormal vestibular group (1.95±1.70)(t=5.305, P<0.01). There were significant differences in the DHI-item 13 among the 3 groups (F=16.402, P<0.01). There was significant difference in VAS scores among the 3 groups (P<0.05). However, the t-test analysis showed that there was significant difference between the BPPV group and the vestibular abnormal group (P<0.05), while there was no significant difference between the BPPV group and the vestibular normal group (P>0.05).
 Conclusion: DHI-2 and DHI-item 13 should be included in the inquiry of disease history at the time of first diagnosis, which can be used to identify patients with BPPV quickly and effectively.

Comparison of chirp versus click and tone pip stimulation for cervical vestibular evoked myogenic potentials.

The current study explored differences among cervical vestibular evoked myogenic potentials (cVEMP) that were evoked by CE-chirp and click and tone pip in healthy controls, and tried to explain the differences of cVEMP between the three of them. Thirty normal volunteers were used as subjects for CE-chirp and click and tone-pip (Blackman pip) stimuli. The latency of P1, N1, peak-to-peak P1-N1 amplitude, and cVEMP interaural difference were obtained and analyzed. The response rates of cVEMP were 93 % for click and 100 % for both Blackman pip and CE-chirp, respectively. The P1 and N1 latencies of cVEMP evoked by CE-chirp were the shortest, followed by click, with Blackman pip the longest (F = 6,686.852, P < 0.001). All indices of cVEMP evoked by the three stimuli showed no significant difference between the left and right ears or between genders. cVEMP responses were significantly different between the three stimuli. Compared with the currently used stimulus, CE-chirp can evoke cVEMP with shorter latencies and demonstrates increased speed and reliability.

Diagnostic Value of CD34 and CD117 Immunohistochemistry and Megakaryocyte Morphology in Myelodysplastic Syndromes: A Retrospective Case-control Study.

This study evaluated the diagnostic value of CD34 and CD117 immunohistochemistry(IHC) and megakaryocyte morphology in Myelodysplastic syndromes (MDS). In this study, CD34-positive individual cells (Type I) and small clusters (Type II) were observed in most cases (91.2%). Type II CD34-positive was seen in 24 (49%) MDS cases, and positive percentage was higher than in acute myelogenous leukemia (AML) or aplastic anemia (AA). Type II CD117-positive were observed in 44 (89.8%) MDS cases and Type I were observed in 5 (10.2%) MDS. Type II CD117-positive percentage was higher than in AML or AA. Megakaryocyte counts were normal or increased in most MDS cases except one. Although megakaryocyte counts of AML and AA were predominantly decreased, Most MDS patients (81.6%) had abnormal megakaryocyte, whereas almost none of megakaryocyte abnormality was found in AML and AA. In conclusion, combined detection of CD34 and CD117 and observation of megakaryocyte count and morphology are useful for the diagnosis of MDS.

Wogonin protects against bleomycin-induced mouse pulmonary fibrosis via the inhibition of CDK9/p53-mediated cell senescence.

Pulmonary fibrosis (PF) is a fatal interstitial lung disease associated with declining pulmonary function but currently with few effective drugs. Cellular senescence has been implicated in the pathogenesis of PF and could be a potential therapeutic target. Emerging evidence suggests wogonin, the bioactive compound isolated from Scutellaria baicalensis, owns the anti-senescence properties, however, the possible impact of wogonin on PF and the potential mechanisms remain unclear. In this study, a well-established mouse model of PF was utilized which mice were administrated with bleomycin (BLM). Strikingly, wogonin treatment significantly reduced fibrosis deposition in the lung induced by BLM. In vitro, wogonin also suppressed fibrotic markers of cultured epithelial cells stimulated by BLM or hydrogen peroxide. Mechanistic investigation revealed that wogonin attenuated the expressions of DNA damage marker γ-H2AX and senescence-related markers including phosphorylated p53, p21, retinoblastoma protein (pRB), and senescence-associated ß-galactosidase (SA-ß-gal). Moreover, wogonin, as a direct and selective inhibitor of cyclin-dependent kinase 9 (CDK9), exhibited anti-fibrotic capacity by inhibiting CDK9 and p53/p21 signalling. In conclusion, wogonin protects against BLM-induced PF in mice through the inhibition of cell senescence via the regulation of CDK9/p53 and DNA damage pathway. This is the first study to demonstrate the beneficial effect of wogonin on PF, and its implication as a novel candidate for PF therapy.

The efficacy of keverprazan-based quadruple therapy for Helicobacter pylori eradication: a phase III, randomised, double-blind, multicentre trial.

INTRODUCTION: Keverprazan is a novel potassium-competitive acid blocker. The advantages of keverprazan as a potent acid suppressor in Helicobacter pylori eradication have not yet been demonstrated. The aim of this study was to evaluate the efficacy of keverprazan as a component of bismuth quadruple therapy in H. pylori treatment. METHODS: Adult patients with H. pylori infection were enrolled and randomised to take keverprazan (KEV group)- or esomeprazole (ESO group)-quadruple therapy. The regimens contained keverprazan 20 mg or esomeprazole 20 mg, clarithromycin 500 mg, amoxicillin 1000 mg and bismuth potassium citrate 240 mg and were administered twice daily for 14 days. The primary endpoint was the H. pylori eradication rate at 4 weeks after treatment. RESULTS: The full analysis set showed that the H. pylori eradication rates were 87.8% (252/287) and 82.52% (236/286) for the KEV and ESO groups, respectively (difference: 5.29%; 95% confidence interval [CI]: -0.55-11.18). Keverprazan was superior to esomeprazole in terms of eradication rate in the per protocol set (P=0.0382). The eradication rates for patients resistant or non-resistant to clarithromycin were both numerically higher in the KEV group than the ESO group (83.45% vs. 76.98% for clarithromycin-resistance; 92.31% vs. 88.16% for clarithromycin-non-resistance). The incidence of adverse events was similar in the KEV and ESO groups (76.31% vs. 77.62%), with most adverse events (>90%) being mild in severity. No TEAEs led to death in either group. CONCLUSIONS: Keverprazan 20 mg twice daily, used as a component of bismuth quadruple therapy, provided effective H. pylori eradication and was non-inferior to an esomeprazole-based regimen.

Identification of PANoptosis-related subtypes, construction of a prognosis signature, and tumor microenvironment landscape of hepatocellular carcinoma using bioinformatic analysis and experimental verification.

Background: Hepatocellular carcinoma (HCC) is one of the most lethal malignancies worldwide. PANoptosis is a recently unveiled programmed cell death pathway, Nonetheless, the precise implications of PANoptosis within the context of HCC remain incompletely elucidated. Methods: We conducted a comprehensive bioinformatics analysis to evaluate both the expression and mutation patterns of PANoptosis-related genes (PRGs). We categorized HCC into two clusters and identified differentially expressed PANoptosis-related genes (DEPRGs). Next, a PANoptosis risk model was constructed using LASSO and multivariate Cox regression analyses. The relationship between PRGs, risk genes, the risk model, and the immune microenvironment was studies. In addition, drug sensitivity between high- and low-risk groups was examined. The expression profiles of these four risk genes were elucidate by qRT-PCR or immunohistochemical (IHC). Furthermore, the effect of CTSC knock down on HCC cell behavior was verified using in vitro experiments. Results: We constructed a prognostic signature of four DEPRGs (CTSC, CDCA8, G6PD, and CXCL9). Receiver operating characteristic curve analyses underscored the superior prognostic capacity of this signature in assessing the outcomes of HCC patients. Subsequently, patients were stratified based on their risk scores, which revealed that the low-risk group had better prognosis than those in the high-risk group. High-risk group displayed a lower Stromal Score, Immune Score, ESTIMATE score, and higher cancer stem cell content, tumor mutation burden (TMB) values. Furthermore, a correlation was noted between the risk model and the sensitivity to 56 chemotherapeutic agents, as well as immunotherapy efficacy, in patient with. These findings provide valuable guidance for personalized clinical treatment strategies. The qRT-PCR analysis revealed that upregulated expression of CTSC, CDCA8, and G6PD, whereas downregulated expression of CXCL9 in HCC compared with adjacent tumor tissue and normal liver cell lines. The knockdown of CTSC significantly reduced both HCC cell proliferation and migration. Conclusion: Our study underscores the promise of PANoptosis-based molecular clustering and prognostic signatures in predicting patient survival and discerning the intricacies of the tumor microenvironment within the context of HCC. These insights hold the potential to advance our comprehension of the therapeutic contribution of PANoptosis plays in HCC and pave the way for generating more efficacious treatment strategies.

Genome-Wide Analysis of Family I84 Protease Inhibitor Genes in Three Bivalves Reveals Important Information About the Protein Family's Evolution.

Family I84 serine protease inhibitors are believed to be mollusk specific proteins involved in host defense. The molecular evolution of the family, however, remains to be understood. In this study, the genes of Family I84 protease inhibitors in 3 bivalves, Crassostrea gigas, Crassostrea virginica and Tegillarca granosa, were analyzed at the genomic level. A total of 66 Family I84 genes (22 in C. gigas, 28 in C. virginica and 16 in T. granosa) were identified from the 3 species. They distributed unevenly in the genomes involving 4 chromosomes in C. gigas and 5 chromosomes in C. virginica and T. granosa and some genes were tandemly duplicated. Most genes had 3 exons with 12 genes having 4 exons and 1 gene having 2 exons. All genes but 1 from C. gigas and 1 from T. granosa encoded peptides with a signal sequence at the N-terminus, and the properties of the predicted mature molecules were similar. Four conserved motifs were identified in the 66 amino acid sequences. Collinear analysis revealed higher collinearity between the 2 oyster species in general genes and in Family I84 genes. Phylogenetic analysis of the 66 genes with those previously reported from 3 other bivalves and 1 gastropod showed that Family I84 protease inhibitor genes from the same species tended to be grouped together in terminal branches of the constructed Maximum likelihood tree, but most internal nodes were poorly supported by the bootstrap values. In addition, differences in expression patterns between the genes of a same species were observed in the developmental stages and tissues of C. gigas and T. granosa. Moreover, the co-expression of genes within Family I84 and Family I84 genes with non-Family I84 were also detected in C. gigas and T. granosa. These results suggested that Family I84 protease inhibitor genes evolved by active duplications and structural and functional diversifications after the speciation of related mollusks, and the diversified protease inhibitor family was likely multifunctional.

Efficacy and Safety of Keverprazan Compared With Lansoprazole in the Treatment of Duodenal Ulcer: A Phase III, Randomized, Double-Blind, Multicenter Trial.

INTRODUCTION: Keverprazan is a novel potassium-competitive acid blocker for the treatment of acid-related disorders requiring potent acid inhibition. This study aimed to establish the noninferiority of keverprazan to lansoprazole in the treatment of patients with duodenal ulcer (DU). METHODS: In this phase III, double-blind, multicenter study, 360 Chinese patients with endoscopically confirmed active DU were randomized 1:1 to take either keverprazan (20 mg) or lansoprazole (30 mg) treatment for up to 6 weeks. The primary end point was DU healing rate at week 6. The secondary end point was DU healing rate at week 4. Symptom improvement and safety were also assessed. RESULTS: Based on the full analysis set, the cumulative healing rates at week 6 were 94.4% (170/180) and 93.3% (166/178) for keverprazan and lansoprazole, respectively (difference: 1.2%; 95% confidence intervel: -4.0%-6.5%). At week 4, the respective healing rates were 83.9% (151/180) and 80.3% (143/178). In the per protocol set, the 6-week healing rates in keverprazan and lansoprazole groups were 98.2% (163/166) and 97.6% (163/167), respectively (difference: 0.6%; 95% confidence intervel: -3.1%-4.4%); the 4-week healing rates were respectively 86.8% (144/166) and 85.6% (143/167). Keverprazan was noninferior to lansoprazole in DU healing after the treatment for 4 and 6 weeks. The incidence of treatment-emergent adverse events was comparable among groups. DISCUSSION: Keverprazan 20 mg had a good safety profile and was noninferior to lansoprazole 30 mg once daily for DU healing.

Molecular characterization of two CuZn-SOD family proteins in the Pacific oyster Crassostrea gigas.

Superoxide dismutases (SOD) are multifamily antioxidant enzymes, playing an important role in the defense against oxidative stress in all organisms. Genomic information indicated the presence of genetic diversification of the copper and zinc SOD (CuZn-SOD) family in oysters. In the present research, we characterized two CuZn-SOD family proteins, Cg-CuZn-SOD and Cg-dominin3, in the Pacific oyster Crassostrea gigas using comprehensive sequence analyses, recombinant proteins and site-directed mutagenesis, and observations of gene expression in larval and adult oysters. We found that Cg-CuZn-SOD possessed sequence and structural elements conserved in a CuZn-SOD molecule and the recombinant protein was confirmed empirically to have the SOD enzyme activity. In contrast, Cg-dominin3 lacked five of the seven residues essential for the conformation of SOD active center and the recombinant protein did not have the enzyme activity. However, recombinant Cg-dominin3 showed strong binding activities toward zinc and copper ions. Substitutions of five conserved His residues in the active center demolished the SOD activity but enhanced the metal binding capacity in Cg-CuZn-SOD. On the other hand, reinstallation of the five His residues that were assumed to be activity essential and lost in evolution did not restore the SOD enzyme activity in Cg-dominin3. Additionally, the coding genes of the two proteins exhibited different patterns of expression during larval development and in adult oyster in response to zinc challenges. These results have led to the discovery of the first cytoplasmic CuZn-SOD molecule and the confirmation of molecular diversification of extracellular CuZn-SOD homologs in oysters.

Genome Wide Identification and Expression Profiling Indicate Expansion of Family I84 Protease Inhibitor via Gene Tandem Duplication and Divergence in Razor Clam Sinonovacula constricta.

Family I84 protease inhibitors represent a novel family in the MEROPS peptidase database and are likely unique for molluscan host defense. Two Family I84 members, scSI-1 and scSI-2, were reported from the razor clam Sinonovacula constricta in a previous research. In the present study, 12 additional genes, named scSI-3 to scSI-14, were identified via genome wide sequence analyses. Among them, 10 genes were predicted to have a signal sequence, but one (scSI-7) was not. Besides, one sequence (scSI-14) was likely to encode a prematurely terminated peptide. The predicted mature peptides shared characteristics including 12 conserved cysteine residues, isoelectric points of 4.98 to 6.11, and molecular weights of 7.1 to 9.3 kDa with previously reported family members. Four motifs were characterized in 13 predicted mature peptides (with exception of scSI-14), which shared two to four conserved cysteine residues, are possibly to form two functional domain comprised 6 cysteine residues, respectively. At genomic level, all the 14 razor clam Family I84 genes were organized into 3 exons and 2 introns; 13 of them clustered in 3 regions of 100 kb on 3 separate chromosomes, suggesting tandem duplications of related genes. The promoter region of all the 14 genes was predicted to share some transcription factor binding sites, in particular those responsive to pathological and physiological stimuli, but no shared motifs were identified. Analyses also revealed differences in expression patterns among the genes. One gene in a tandem duplicated gene pairs usually showed a higher expression level than the other whereas non-tandem duplicated genes exhibited a higher degree of correlation in expression level. In addition, 8 of the 14 genes demonstrated higher level of expression in Vibrio tolerant clams than in non-tolerant clams following challenges with Vibrio parahaemolyticus. These results generated important information about the evolution of Family I84 protease inhibitors in S. constricta.

Familial Temperature-Sensitive Auditory Neuropathy: Distinctive Clinical Courses Caused by Variants of the OTOF Gene.

Objective: To investigate the clinical course and genetic etiology of familial temperature-sensitive auditory neuropathy (TSAN), which is a very rare subtype of auditory neuropathy (AN) that involves an elevation of hearing thresholds due to an increase in the core body temperature, and to evaluate the genotype-phenotype correlations in a family with TSAN. Methods: Six members of a non-consanguineous Chinese family, including four siblings complaining of communication difficulties when febrile, were enrolled in this study. The clinical and audiological profiles of the four siblings were fully evaluated during both febrile and afebrile episodes, and the genetic etiology of hearing loss (HL) was explored using next-generation sequencing (NGS) technology. Their parents, who had no complaints of fluctuating HL due to body temperature variation, were enrolled for the genetics portion only. Results: Audiological tests during the patients' febrile episodes met the classical diagnostic criteria for AN, including mild HL, poor speech discrimination, preserved cochlear microphonics (CMs), and absent auditory brainstem responses (ABRs). Importantly, unlike the pattern observed in previously reported cases of TSAN, the ABRs and electrocochleography (ECochG) signals of our patients improved to normal during afebrile periods. Genetic analysis identified a compound heterozygous variant of the OTOF gene (which encodes the otoferlin protein), including one previously reported pathogenic variant, c.5098G > C (p.Glu1700Gln), and one novel variant, c.4882C > A (p.Pro1628Thr). Neither of the identified variants affected the C2 domains related to the main function of otoferlin. Both variants faithfully cosegregated with TSAN within the pedigree, suggesting that OTOF is the causative gene of the autosomal recessive trait segregation in this family. Conclusion: The presence of CMs with absent (or markedly abnormal) ABRs is a reliable criterion for diagnosing AN. The severity of the phenotype caused by dysfunctional neurotransmitter release in TSAN may reflect variants that alter the C2 domains of otoferlin. The observations from this study enrich the current understanding of the phenotype and genotype of TSAN and may lay a foundation for further research on its pathogenesis.

Identification of HSF1 Target Genes Involved in Thermal Stress in the Pacific Oyster Crassostrea gigas by ChIP-seq.

The Pacific oyster Crassostrea gigas, a commercially important species inhabiting the intertidal zone, facing enormous temperature fluctuations. Therefore, it is important to identify candidate genes and key regulatory relationships associated with thermal tolerance, which can aid the molecular breeding of oysters. Heat shock transcription factor 1 (HSF1) plays an important role in the thermal stress resistance. However, the regulatory relationship between the expansion of heat shock protein (HSP) HSP 70 and HSF1 is not yet clear in C. gigas. In this study, we analyzed genes regulated by HSF1 in response to heat shock by chromatin immunoprecipitation followed by sequencing (ChIP-seq), determined the expression patterns of target genes by qRT-PCR, and validated the regulatory relationship between one HSP70 and HSF1. We found 916 peaks corresponding to HSF1 binding sites, and these peaks were annotated to the nearest genes. In Gene Ontology analysis, HSF1 target genes were related to signal transduction, energy production, and response to biotic stimulus. Four HSP70 genes, two HSP40 genes, and one small HSP gene exhibited binding to HSF1. One HSP70 with a binding site in the promoter region was validated to be regulated by HSF1 under heat shock. These results provide a basis for future studies aimed at determining the mechanisms underlying thermal tolerance and provide insights into gene regulation in the Pacific oyster.

Regulation Between HSF1 Isoforms and HSPs Contributes to the Variation in Thermal Tolerance Between Two Oyster Congeners.

Heat shock transcription factor 1 (HSF1) plays an important role in regulating heat shock, which can activate heat shock proteins (HSPs). HSPs can protect organisms from thermal stress. Oysters in the intertidal zone can tolerate thermal stress. The Pacific oyster (Crassostrea gigas gigas) and Fujian oyster (C. gigas angulata)-allopatric subspecies with distinct thermal tolerances-make good study specimens for analyzing and comparing thermal stress regulation. We cloned and compared HSF1 isoforms, which is highly expressed under heat shock conditions in the two subspecies. The results revealed that two isoforms (HSF1a and HSF1d) respond to heat shock in both Pacific and Fujian oysters, and different heat shock conditions led to various combinations of isoforms. Subcellular localization showed that isoforms gathered in the nucleus when exposed to heat shock. The co-immunoprecipitation revealed that HSF1d can be a dimer. In addition, we selected HSPs that are expressed under the heat shock response, according to the RNA-seq and proteomic analyses. For the HSPs, we analyzed the coding part and the promoter sequences. The result showed that the domains of HSPs are conserved in two subspecies, but the promoters are significantly different. The Dual-Luciferase assay showed that the induced expression isoform HSF1d had the highest activity in C. gigas gigas, while the constitutively-expressed HSF1a was most active in C. gigas angulata. In addition, variation in the level of HSP promoters appeared to be correlated with gene expression. We argue that this gene is regulated based on the different expression levels between the two subspecies' responses to heat shock. In summary, various stress conditions can yield different HSF1 isoforms and respond to heat shock in both oyster subspecies. Differences in how the isoforms and promoter are activated may contribute to their differential expressions. Overall, the results comparing C. gigas gigas and C. gigas angulata suggest that these isoforms have a regulatory relationship under heat shock, providing valuable information on the thermal tolerance mechanism in these commercially important oyster species.

[Association of CXCR4 and SDF-1alpha with organ-specific metastasis of nasopharyngeal carcinoma].

OBJECTIVE: To explore the role of CXCR4/SDF-1alpha axis in organ-specific metastasis of nasopharyngeal carcinoma (NPC) by assessment of CXCR4 expression in NPC cells and SDF-1alpha expression in distant target organs of NPC. METHODS: Thirty patients with NPC and fifteen normal subjects were recruited in this study. The expressions of CXCR4 in NPC and normal cases were identified by RT-PCR and immunohistochemistry (IHC), then the relationship between CXCR4 expression and clinicopathological factors was analyzed. IHC was also used to analyze the SDF-1alpha protein expression in normal cervical lymph nodes (including normal superior and inferior deep cervical lymph nodes), bone marrow, lung, liver, kidney and colon tissues of NPC patients (5 cases/each group). RESULTS: The relative expression level of CXCR4 mRNA in NPC (0.71 +/- 0.22) was significantly higher than that of normal nasopharynx tissues (0.14 +/- 0.07, F = 27.94, P < 0.05). The relative expression level of CXCR4 protein in NPC (1.58 +/- 0.59) was significantly higher than that of normal nasopharynx tissues (0.51 +/- 0.22, F = 17.75, P < 0.05). The high expression levels of CXCR4 mRNA and protein in NPC were closely related to clinical stage, cervical lymph node metastasis and cancer cell differentiation (P < 0.05). SDF-1alpha protein was strongly expressed in normal superior deep cervical lymph nodes, bone marrow, lung and liver (2.35 +/- 0.67), but absent or very poor expression in inferior deep cervical lymph nodes, kidney and colon tissues (0.68 +/- 0.23), and the differences between them were statistically significant (t = 10.13, P < 0.01). CONCLUSION: CXCR4 is closely correlated to metastasis of nasopharyngeal carcinoma. CXCR4/SDF-1alpha axis may play an important role in organ-specific metastasis of NPC.

Divergence and plasticity shape adaptive potential of the Pacific oyster.

The interplay between divergence and phenotypic plasticity is critical to our understanding of a species' adaptive potential under rapid climate changes. We investigated divergence and plasticity in natural populations of the Pacific oyster Crassostrea gigas with a congeneric oyster Crassostrea angulata from southern China used as an outgroup. Genome re-sequencing of 371 oysters revealed unexpected genetic divergence in a small area that coincided with phenotypic divergence in growth, physiology, heat tolerance and gene expression across environmental gradients. These findings suggest that selection and local adaptation are pervasive and, together with limited gene flow, influence population structure. Genes showing sequence differentiation between populations also diverged in transcriptional response to heat stress. Plasticity in gene expression is positively correlated with evolved divergence, indicating that plasticity is adaptive and favoured by organisms under dynamic environments. Divergence in heat tolerance-partly through acetylation-mediated energy depression-implies differentiation in adaptive potential. Trade-offs between growth and survival may play an important role in local adaptation of oysters and other marine invertebrates.