Potential role of host autophagy in Clonorchis sinensis infection.

An in vivo mouse model of Clonorchis sinensis (C. sinensis) infection with or without the administration of autophagy inhibitor chloroquine (CQ) stimulation was established to assess the possible involvement of autophagic response during C. sinensis infection. Abnormal liver function was observed at 4, 6, and 8 weeks post-infection, as indicated by elevated levels of ALT/GPT, AST/GOT, TBIL, and α-SMA in the infected groups. These findings indicated that C. sinensis infection activated autophagy, as shown by a decreased LC3II/I ratio and accumulated P62 expression in infected mice. Interestingly, CQ administration exhibited dual and opposing effects during the infection. In the early stage of infection, the engagement of CQ appeared to mitigate symptoms by reducing inflammation and fibrotic responses. However, in the later stage of infection, CQ might contribute to parasite survival by evading autophagic targeting, thereby exacerbating hepatic impairment and worsening liver fibrosis. Autophagy in liver was suppressed throughout the infection. These observations attested that C. sinensis infection triggered autophagy, and highlighted a complex role for CQ, with both protective and detrimental effects, in the in vivo process of C. sinensis infection.

Related Factors with Vascular Dementia: A Two-Sample Mendelian Randomization Study.

Pathogenesis of vascular dementia (VD) is still unclear, there are currently no effective prevention and treatment methods. We applied Mendelian randomization (MR) using summary statistics from large-scale GWAS of metabolites and VD to reveal the causal effect of metabolites on the VD. One set of genetics instrument was used for analysis, derived from publicly available genetic summary data. Which was 32 single-nucleotide polymorphisms robustly associated with metabolites. Inverse-variance weighted, weighted median method, MR-Egger regression, and MR Pleiotropy RESidual Sum and Outlier test were used for MR analyses. Strong evidence for a positive effect of metabolites, which means N6-threonylcarbamoyladenosine (t6A) on VD was found in inverse-variance weighted (odds ratios [OR]: 0.667, 95% confidence interval [CI]: 0.548-0.812, p < 0.001), MR-Egger (OR: 0.647, 95% CI: 0.458-0.913, p = 0.019), and weighted median (OR: 0.650, 95% CI: 0.466-0.908, p = 0.012). The MR analysis indicated that metabolites (t6A) may be causally associated with a positive effect on VD.

Natural history of spinal cord compression stage AFMS3 in infants with achondroplasia: retrospective cohort study.

BACKGROUND AND OBJECTIVE: Foramen magnum stenosis (FMS) is a common, serious complication of achondroplasia in infancy and associated with sudden infant death. The Achondroplasia Foramen Magnum Score (AFMS; 0-4) is used to classify the severity of stenosis to inform appropriate neurosurgical management. Infants with AFMS4 are referred for neurosurgery, while well children with AFMS3 undergo repeat MRI routinely after 12 months.As the natural history of children with AFMS3 is currently unclear, the objective was to review follow-up MRI scans of infants initially classified as AFMS3 to define more clearly the evolution of this degree of stenosis. DESIGN: This retrospective cohort study, from two tertiary centres, included infants with a confirmed diagnosis of achondroplasia and AFMS3 on initial MRI who subsequently underwent repeat MRI or proceeded straight to neurosurgery. RESULTS: Twenty-two cases satisfied the inclusion criteria. Mean age in months was 6.23 (SD±3.82) and 17.95 (SD±7.68) at baseline and follow-up scans, respectively. Follow-up MRI showed no change in 23% (N=5), improvement in 36% (N=8) to either AFMS1 (N=5) or AFMS2 (N=3). There was progression in 41% to AFMS4 (N=8). One case had neurosurgey without follow-up MRI (N=1). CONCLUSIONS: These results support MRI screening for FMS in infants with achondroplasia. Furthermore, infants with AFMS3 should undergo follow-up MRI as over 40% progress prompting neurosurgical intervention. There is currently no consensus on frequency or timing of screening for AFMS3 in achondroplasia; however, we suggest that guidance for follow-up imaging is modified to 6 months to detect progression earlier in this at-risk cohort.

Reversible Sterilization of Channel Catfish via Overexpression of Glutamic Acid Decarboxylase Gene.

The confinement of transgenic fish is essential to prevent their escape and reproduction in natural ecosystems. Reversible transgenic sterilization is a promising approach to control the reproduction of transgenic fish. Therefore, the present study was conducted to develop a reversibly sterile channel catfish (Ictalurus punctatus) via the transgenic overexpression of the goldfish (Carassius auratus) glutamic acid decarboxylase (GAD) gene driven by the common carp (Cyprinus carpio) ß-actin promoter to disrupt normal gamma-aminobutyric acid (GABA) regulation. Three generations of GAD-transgenic fish were produced. All studied generations showed repressed reproductive performance; however, this was not always statistically significant. In F1, 5.4% of the transgenic fish showed a sexual maturity score ≥ 4 (maximum = 5) at five years of age, which was lower (p = 0.07) than that of the control group (16.8%). In the spawning experiments conducted on F1 transgenic fish at six and nine years of age, 45.5% and 20.0% of fish spawned naturally, representing lower values (p = 0.09 and 0.12, respectively) than the percentages in the sibling control fish of the same age (83.3% and 66.7%, respectively). Four of six pairs of the putative infertile six-year-old fish spawned successfully after luteinizing hormone-releasing hormone analog (LHRHa) therapy. Similar outcomes were noted in the three-year-old F2 fish, with a lower spawning percentage in transgenic fish (20.0%) than in the control (66.7%). In one-year-old F2-generation transgenic fish, the observed mean serum gonadotropin-releasing hormone (GnRH) levels were 9.23 ± 2.49 and 8.14 ± 2.21 ng/mL for the females and males, respectively. In the control fish, the mean levels of GnRH were 11.04 ± 4.06 and 9.03 ± 2.36 ng/mL for the females and males, respectively, which did not differ significantly from the control (p = 0.15 and 0.27 for females and males, respectively). There was no significant difference in the estradiol levels of the female transgenic and non-transgenic fish in the one- and four-year-old F2-generation fish. The four-year-old F2-generation male transgenic fish exhibited significantly (p < 0.05) lower levels of GnRH and testosterone than the control fish. In conclusion, while overexpressing GAD repressed the reproductive abilities of channel catfish, it did not completely sterilize transgenic fish. The sterilization rate might be improved through selection in future generations.

Advancing aquaculture: Production of xenogenic catfish by transplanting blue catfish (Ictalurus furcatus) and channel catfish (I. punctatus) stem cells into white catfish (Ameiurus catus) triploid fry.

Xenogenesis has been recognized as a prospective method for producing channel catfish, Ictalurus punctatus ♀ × blue catfish, I. furcatus ♂ hybrids. The xenogenesis procedure can be achieved by transplanting undifferentiated stem cells derived from a donor fish into a sterile recipient. Xenogenesis for hybrid catfish embryo production has been accomplished using triploid channel catfish as a surrogate. However, having a surrogate species with a shorter maturation period, like white catfish (Ameiurus catus), would result in reduced feed costs, labor costs, and smaller body size requirements, making it a more suitable species for commercial applications where space is limited, and as a model species. Hence, the present study was conducted to assess the effectiveness of triploid white catfish as a surrogate species to transplant blue catfish stem cells (BSCs) and channel catfish stem cells (CSCs). Triploid white catfish fry were injected with either BSCs or CSCs labeled with PKH 26 fluorescence dye from 0 to 12 days post hatch (DPH). No significant differences in weight and length of fry were detected among BSCs and CSCs injection times (0 to 12 DPH) when fry were sampled at 45 and 90 DPH (P > 0.05). The highest survival was reported when fry were injected between 4.0 to 5.5 DPH (≥ 81.2%). At 45 and 90 DPH, cell and cluster area increased for recipients injected from 0 to 5.2 DPH, and the highest cluster area values were reported between 4.0 to 5.2 DPH. Thereafter, fluorescent cell and cluster area in the host declined with no further decrease after 10 DPH. At 45 DPH, the highest percentage of xenogens were detected when fry were injected with BSCs between 4.0 to 5.0 and CSCs between 3.0 to 5.0 DPH. At 90 DPH, the highest number of xenogens were detected from 4.0 to 6.0 DPH when injected with either BSCs or CSCs. The current study demonstrated the suitability of white catfish as a surrogate species when BSCs and CSCs were transplanted into triploid white catfish between 4.0 to 6.0 DPH (27.4 ± 0.4°C). Overall, these findings allow enhanced efficiency of commercializing xenogenic catfish carrying gametes of either blue catfish or channel catfish.

Serum 25-hydroxyvitamin D level and erectile dysfunction: a causal relationship? Findings from a two-sample Mendelian randomization study.

Background: Serum 25-hydroxyvitamin D level is associated with erectile dysfunction (ED) in observational studies. However, whether there is a causal association between them remains uncertain. Objective: Conduct a two-sample Mendelian randomization (MR) analysis to investigate the causal effect between serum 25-hydroxyvitamin D level and ED risk. Method: Genome-wide association study (GWAS) data of serum 25-hydroxyvitamin D levels comprising 6,896,093 single nucleotide polymorphisms (SNP) from 496,949 people of European ancestry were regarded as exposure for the MR analysis. Additional GWAS data involving 9,310,196 SNPs of 6,175 European ED cases and 217,630 controls were used as outcome data. The MR-Egger, inverse variance weighted (IVW) method, weighted median, simple mode, and weighted mode were employed to evaluate causal effects, among which IVW was the primary MR analysis method. The stability of the MR analysis results was confirmed by a heterogeneity test, a horizontal pleiotropy test, and the leave-one-out method. Result: There were 103 SNPs utilized as instrumental variables (p < 5 × 10-8). The results of MR analysis showed no causal effects of serum 25(OH) D concentration on ED risks (IVW; OR = 0.9516, 95% CI = 0.7994 to 1.1328, p = 0.5772). There was no heterogeneity and pleiotropy in the statistical models. Conclusion: The present MR study did not support a causal association for genetically predicted serum 25-hydroxyvitamin D concentration in the risk of ED in individuals of European descent.

Transcriptome analysis revealed potential mechanisms of channel catfish growth advantage over blue catfish in a tank culture environment.

Channel catfish (Ictalurus punctatus) and blue catfish (Ictalurus furcatus) are two economically important freshwater aquaculture species in the United States, with channel catfish contributing to nearly half of the country's aquaculture production. While differences in economic traits such as growth rate and disease resistance have been noted, the extent of transcriptomic variance across various tissues between these species remains largely unexplored. The hybridization of female channel catfish with male blue catfish has led to the development of superior hybrid catfish breeds that exhibit enhanced growth rates and improved disease resistance, which dominate more than half of the total US catfish production. While hybrid catfish have significant growth advantages in earthen ponds, channel catfish were reported to grow faster in tank culture environments. In this study, we confirmed channel fish's superiority in growth over blue catfish in 60-L tanks at 10.8 months of age (30.3 g and 11.6 g in this study, respectively; p < 0.001). In addition, we conducted RNA sequencing experiments and established transcriptomic resources for the heart, liver, intestine, mucus, and muscle of both species. The number of expressed genes varied across tissues, ranging from 5,036 in the muscle to over 20,000 in the mucus. Gene Ontology analysis has revealed the functional specificity of differentially expressed genes within their respective tissues, with significant pathway enrichment in metabolic pathways, immune activity, and stress responses. Noteworthy tissue-specific marker genes, including lrrc10, fabp2, myog, pth1a, hspa9, cyp21a2, agt, and ngtb, have been identified. This transcriptome resource is poised to support future investigations into the molecular mechanisms underlying environment-dependent heterosis and advance genetic breeding efforts of hybrid catfish.

Association of the Achondroplasia Foramen Magnum Score and intraoperative neuromonitoring.

INTRODUCTION: Foramen magnum stenosis in achondroplasia carries a risk of sudden death. A proportion of these patients benefit from foramen magnum decompression (FMD). The Achondroplasia Foramen Magnum Score (AFMS) was developed to stratify those most at risk. We hypothesise that this score may be reflected in neurophysiological findings. METHODS: Patients with achondroplasia who had undergone FMD (n=20) were retrospectively grouped into AFMS 2, 3 and 4. Amplitude from tibialis anterior (TA) and the percentage change in somatosensory evoked potential (SSEP) latency after FMD were reported. RESULTS: Baseline motor evoked potential amplitudes for patients with AFMS=4 were significantly lower left (p=0.0017 and p=0.02 for right and left TA, respectively) compared with AFMS grades 2 and 3. Median reduction (% change) in SSEP latency (ms) after surgery was not significantly different in any of the patients. CONCLUSIONS: This short report cross-references AFMS to intraoperative neuromonitoring. Baseline amplitudes were noticeably lower in the most severe AFMS group. This observation supports the notion that AFMS can help risk stratify patients and aid in surgical selection.

From heterosis to outbreeding depression: genotype-by-environment interaction shifts hybrid fitness in opposite directions.

In F1 hybrids, phenotypic values are expected to be near the parental means under additive effects or close to one parent under dominance. However, F1 traits can fall outside the parental range, and outbreeding depression occurs when inferior fitness is observed in hybrids. Another possible outcome is heterosis, a phenomenon that interspecific hybrids or intraspecific crossbred F1s exhibit improved fitness compared to both parental species or strains. As an application of heterosis, hybrids between channel catfish females and blue catfish males are superior in feed conversion efficiency, carcass yield, and harvestability. Over 20 years of hybrid catfish production in experimental settings and farming practices generated abundant phenotypic data, making it an ideal system to investigate heterosis. In this study, we characterized fitness in terms of growth and survival longitudinally, revealing environment-dependent heterosis. In ponds, hybrids outgrow both parents due to an extra rapid growth phase of 2-4 months in year 2. This bimodal growth pattern is unique to F1 hybrids in pond culture environments only. In sharp contrast, the same genetic types cultured in tanks display outbreeding depression, where hybrids perform poorly, while channel catfish demonstrate superiority in growth throughout development. Our findings represent the first example, known to the authors, of opposite fitness shifts in response to environmental changes in interspecific vertebrate hybrids, suggesting a broader fitness landscape for F1 hybrids. Future genomic studies based on this experiment will help understand genome-environment interaction in shaping the F1 progeny fitness in the scenario of environment-dependent heterosis and outbreeding depression.

Corrigendum: A comprehensive analysis of HAVCR1 as a prognostic and diagnostic marker for pan-cancer.

[This corrects the article DOI: 10.3389/fgene.2022.904114.].

Natural products and their derivatives alleviating cerebral white matter lesions.

White matter lesions (WMLs), characterized by focal demyelination or myelination disorders, are commonly present in cerebral small vessel disease and various neurological diseases. Multiple etiologies lead to WMLs. However, there is no specific therapy or effective drugs for relieving WMLs. Natural products and their derivatives originate from bacterial, fungal, plant, and marine animal sources, many of which have multiple therapeutic targets. Compared to single target compounds, natural products and their derivatives are promising to be developed as better drugs to attenuate WMLs. Thus, this review attempts to summarize the status of natural products and their derivatives (2010-to date) alleviating cerebral white matter lesions for the discovery of new drugs.

One-step knock-in of two antimicrobial peptide transgenes at multiple loci of catfish by CRISPR/Cas9-mediated multiplex genome engineering.

CRISPR/Cas9-mediated multiplex genome editing (MGE) conventionally uses multiple single-guide RNAs (sgRNAs) for gene-targeted mutagenesis via the non-homologous end joining (NHEJ) pathway. MGE has been proven to be highly efficient for functional gene disruption/knockout (KO) at multiple loci in mammalian cells or organisms. However, in the absence of a DNA donor, this approach is limited to small indels without transgene integration. Here, we establish the linear double-stranded DNA (dsDNA) and double-cut plasmid (dcPlasmid) combination-assisted MGE in channel catfish (Ictalurus punctatus), allowing combinational deletion mutagenesis and transgene knock-in (KI) at multiple sites through NHEJ/homology-directed repair (HDR) pathway in parallel. In this study, we used single-sgRNA-based genome editing (ssGE) and multi-sgRNA-based MGE (msMGE) to replace the luteinizing hormone (lh) and melanocortin-4 receptor (mc4r) genes with the cathelicidin (As-Cath) transgene and the myostatin (two target sites: mstn1, mstn2) gene with the cecropin (Cec) transgene, respectively. A total of 9000 embryos were microinjected from three families, and 1004 live fingerlings were generated and analyzed. There was no significant difference in hatchability (all P > 0.05) and fry survival (all P > 0.05) between ssGE and msMGE. Compared to ssGE, CRISPR/Cas9-mediated msMGE assisted by the mixture of dsDNA and dcPlasmid donors yielded a higher knock-in (KI) efficiency of As-Cath (19.93 %, [59/296] vs. 12.96 %, [45/347]; P = 0.018) and Cec (22.97 %, [68/296] vs. 10.80 %, [39/361]; P = 0.003) transgenes, respectively. The msMGE strategy can be used to generate transgenic fish carrying two transgenes at multiple loci. In addition, double and quadruple mutant individuals can be produced with high efficiency (36.3 % âˆ¼ 71.1 %) in one-step microinjection. In conclusion, we demonstrated that the CRISPR/Cas9-mediated msMGE allows the one-step generation of simultaneous insertion of the As-Cath and Cec transgenes at four sites, and the simultaneous disruption of the lh, mc4r, mstn1 and mstn2 alleles. This msMGE system, aided by the mixture donors, promises to pioneer a new dimension in the drive and selection of multiple designated traits in other non-model organisms.

Effects of co-infection with Clonorchis sinensis on T cell exhaustion levels in patients with chronic hepatitis B.

To investigate the effects of co-infection with Clonorchis sinensis (C. sinensis) on T cell exhaustion levels in patients with chronic hepatitis B, we enrolled clinical cases in this study, including the patients with concomitant C. sinensis and HBV infection. In this study, we detected inhibitory receptors and cytokine expression in circulating CD4+ and CD8+ T cells by flow cytometry. PD-1 and TIM-3 expression levels were significantly higher on CD4+ T and CD8+ T cells from co-infected patients than on those from the HBV patients. In addition, CD4+ T cells and CD8+ T cells function were significantly inhibited by C. sinensis and HBV co-infection compared with HBV single infection, secreting lower levels of Interferon gamma (IFN-γ), Interleukin-2 (IL-2), and TNF-α. Our current results suggested that C. sinensis co-infection could exacerbate T cell exhaustion in patients with chronic hepatitis B. PD-1 and TIM-3 could be novel biomarkers for T cell exhaustion in patients with Clonorchis sinensis and chronic hepatitis B co-infection. Furthermore, it may be one possible reason for the weaker response to antiviral therapies and the chronicity of HBV infection in co-infected patients. We must realize the importance of C. sinensis treatment for HBV-infected patients. It might provide useful information for clinical doctors to choose the right treatment plans.

Allergen sensitization study in Dongying, China: An epidemiological study.

BACKGROUND: To explore the relationship between specific immunoglobulin E levels in response to prevalent pollen and food allergens among patients suffering from localized allergic diseases in the Dongying area of China, and to analyze the interconnectivity among these factors. METHODS: This research encompassed allergic patients who visited the Allergy Department of Shengli Oilfield Central Hospital from January 2022 to January 2023. We examined the specific immunoglobulin E levels in the blood of 230 patients utilizing the Fobock platform provided by Jiangsu Haoeubo Company. Statistical analysis was conducted with SPSS 25.0 statistical software. The chi-square test evaluated the relevance of differences in gender and age. A value of P < .05 was considered statistically significant. RESULTS: In this study, eggs emerged as the allergen with the highest number of sensitized individuals, closely followed by dust mite. Conversely, the least sensitized allergen was the cypress tree, closely followed by mango. Notably, male patients exhibited higher sensitivities to cottonwood (P < .05) and egg (P < .001) compared to female patients. Children aged 0 to 10 years showed increased sensitivity to variety of allergens. A significant correlation was observed among different allergens. The top ten allergen pairs with the highest correlation included Birch Tree and Cottonwood (0.88, P < .001), Cottonwood and Pine Tree (0.86, P < .001), Birch Tree and Pine Tree (0.84, P < .001), Pine Tree and Paulownia (0.81, P < .001), Dust Mite and House Dust Mite (0.76, P < .001), Birch Tree and Paulownia (0.73, P < .001), Cashew and Pistachio (0.71, P < .001), Apple and Hazelnut (0.71, P < .001), Cottonwood and Paulownia (0.71, P < .001), and Pine Tree and Ordinary Ragweed (0.70, P < .001). CONCLUSION: This research sheds light on the patterns of allergen sensitization in Dongying, Shandong, highlighting that egg is the most prevalent sensitizing allergen. A notably high correlation was observed between Birch Tree and Cottonwood. This study enhanced the understanding of allergic diseases, explored the causes and mechanisms of allergies, strengthened the management of allergic diseases. Furthermore, it offers valuable insights for the clinical diagnosis and prevention of allergic diseases.

Development of a novel integrated isothermal amplification system for detection of bacteria-spiked blood samples.

Bloodstream infection (BSI) caused by bacteria is highly pathogenic and lethal, and easily develops whole-body inflammatory state. Immediate identification of disease-causing bacteria can improve patient prognosis. Traditional testing methods are not only time-consuming, but such tests are limited to laboratories. Recombinase polymerase amplification combined with lateral flow dipstick (RPA-LFD) holds great promise for rapid nucleic acid detection, but the uncapping operation after amplification easily contaminates laboratories. Therefore, the establishment of a more effective integrated isothermal amplification system has become an urgent problem to be solved. In this study, we designed and fabricated a hermetically sealed integrated isothermal amplification system. Combining with this system, a set of RPA-LFD assays for detecting S. aureus, K. peneumoniae, P. aeruginosa, and H. influenza in BSI were established and evaluated. The whole process could be completed in less than 15 min and the results can be visualized by the naked eye. The developed RPA-LFD assays displayed a good sensitivity, and no cross-reactivity was observed in seven similar bacterial genera. The results obtained with 60 clinical samples indicated that the developed RPA-LFD assays had high specifcity and sensitivity for identifying S. aureus, K. peneumoniae, P. aeruginosa, and H. influenza in BSI. In conclusion, our results showed that the developed RPA-LFD assay is an alternative to existing PCR-based methods for detection of S. aureus, K. peneumoniae, P. aeruginosa, and H. influenza in BSI in primary hospitals.

Simultaneous estimation of gene regulatory network structure and RNA kinetics from single cell gene expression.

Cells respond to environmental and developmental stimuli by remodeling their transcriptomes through regulation of both mRNA transcription and mRNA decay. A central goal of biology is identifying the global set of regulatory relationships between factors that control mRNA production and degradation and their target transcripts and construct a predictive model of gene expression. Regulatory relationships are typically identified using transcriptome measurements and causal inference algorithms. RNA kinetic parameters are determined experimentally by employing run-on or metabolic labeling (e.g. 4-thiouracil) methods that allow transcription and decay rates to be separately measured. Here, we develop a deep learning model, trained with single-cell RNA-seq data, that both infers causal regulatory relationships and estimates RNA kinetic parameters. The resulting in silico model predicts future gene expression states and can be perturbed to simulate the effect of transcription factor changes. We acquired model training data by sequencing the transcriptomes of 175,000 individual Saccharomyces cerevisiae cells that were subject to an external perturbation and continuously sampled over a one hour period. The rate of change for each transcript was calculated on a per-cell basis to estimate RNA velocity. We then trained a deep learning model with transcriptome and RNA velocity data to calculate time-dependent estimates of mRNA production and decay rates. By separating RNA velocity into transcription and decay rates, we show that rapamycin treatment causes existing ribosomal protein transcripts to be rapidly destabilized, while production of new transcripts gradually slows over the course of an hour. The neural network framework we present is designed to explicitly model causal regulatory relationships between transcription factors and their genes, and shows superior performance to existing models on the basis of recovery of known regulatory relationships. We validated the predictive power of the model by perturbing transcription factors in silico and comparing transcriptome-wide effects with experimental data. Our study represents the first step in constructing a complete, predictive, biophysical model of gene expression regulation.

CRISPR/Cas9-mediated knock-in of masu salmon (Oncorhyncus masou) elongase gene in the melanocortin-4 (mc4r) coding region of channel catfish (Ictalurus punctatus) genome.

Channel catfish, Ictalurus punctatus, have limited ability to synthesize Ω-3 fatty acids. The ccßA-msElovl2 transgene containing masu salmon, Oncorhynchus masou, elongase gene driven by the common carp, Cyprinus carpio, ß-actin promoter was inserted into the channel catfish melanocortin-4 receptor (mc4r) gene site using the two-hit two-oligo with plasmid (2H2OP) method. The best performing sgRNA resulted in a knockout mutation rate of 92%, a knock-in rate of 54% and a simultaneous knockout/knock-in rate of 49%. Fish containing both the ccßA-msElovl2 transgene knock-in and mc4r knockout (Elovl2) were 41.8% larger than controls at 6 months post-hatch (p = 0.005). Mean eicosapentaenoic acid (EPA, C20:5n-3) levels in Elov2 mutants and mc4r knockout mutants (MC4R) were 121.6% and 94.1% higher than in controls, respectively (p = 0.045; p = 0.025). Observed mean docosahexaenoic acid (DHA, C22:6n-3) and total EPA + DHA content was 32.8% and 45.1% higher, respectively, in Elovl2 transgenic channel catfish than controls (p = 0.368; p = 0.025). To our knowledge this is the first example of genome engineering to simultaneously target transgenesis and knock-out a gene in a commercially important aquaculture species for multiple improved performance traits. With a high transgene integration rate, improved growth, and higher omega-3 fatty acid content, the use of Elovl2 transgenic channel catfish appears beneficial for application on commercial farms.

Effect of Lacking ZKSCAN3 on Autophagy, Lysosomal Biogenesis and Senescence.

Transcription factors can affect autophagy activity by promoting or inhibiting the expression of autophagic and lysosomal genes. As a member of the zinc finger family DNA-binding proteins, ZKSCAN3 has been reported to function as a transcriptional repressor of autophagy, silencing of which can induce autophagy and promote lysosomal biogenesis in cancer cells. However, studies in Zkscan3 knockout mice showed that the deficiency of ZKSCAN3 did not induce autophagy or increase lysosomal biogenesis. In order to further explore the role of ZKSCAN3 in the transcriptional regulation of autophagic genes in human cancer and non-cancer cells, we generated ZKSCAN3 knockout HK-2 (non-cancer) and Hela (cancer) cells via the CRISPR/Cas9 system and analyzed the differences in gene expression between ZKSCAN3 deleted cells and non-deleted cells through fluorescence quantitative PCR, western blot and transcriptome sequencing, with special attention to the differences in expression of autophagic and lysosomal genes. We found that ZKSCAN3 may be a cancer-related gene involved in cancer progression, but not an essential transcriptional repressor of autophagic or lysosomal genes, as the lacking of ZKSCAN3 cannot significantly promote the expression of autophagic and lysosomal genes.

MiR-455-5p suppresses PDZK1IP1 to promote the motility of oral squamous cell carcinoma and accelerate clinical cancer invasion by regulating partial epithelial-to-mesenchymal transition.

BACKGROUND: Lymph node and distant metastasis contribute to poor outcomes in patients with oral squamous cell carcinoma (OSCC). The mechanisms regulating cancer migration and invasion play a key role in OSCC. METHODS: We determined migration and invasion ability of OSCC by wound-healing assay, two-chamber transwell invasion assay and cell mobility tracking and evaluated tumor metastasis in vivo. Western blot (WB), qRT-PCR, RNA-seq, dual-luciferase reporter assays and nuclear/cytoplasmic fractionation were performed to investigate the potential mechanism. Immunohistochimical (IHC) staining determined vimentin and PDZK1IP1 expression in OSCC tissues. RESULTS AND CONCLUSION: In this study, we determined that miR-455-5p was associated with lymph node metastasis and clinical invasion, leading to poor outcomes in patients with OSCC. MiR-455-5p promoted oral cancer cell migration and invasion and induced epithelial-to-mesenchymal transition (EMT). We also identified a new biomarker, PDZK1IP1 (MAP17), that was targeted by miR-455-5p. PDZK1IP1 knockdown led to migration, metastasis, EMT, and increased transforming growth factor-ß signaling in OSCC. In addition, miR-455-5p overexpression and PDZK1IP1 inhibition promoted collective OSCC cell migration. According to data from the Cancer Genome Atlas database and the NCKU-OrCA-40TN data set, miR-455-5p and PDZK1IP1 are positively and negatively correlated, respectively, with partial EMT score. High miR-455-5p expression was associated with high vimentin levels and low MAP17 H-scores. The patients with low MAP17 expression had higher rates of disease recurrence than did patients with high MAP17 expression, especially for patients with clinical invasion risk factors and low MAP17 expression. These results suggest that miR-455-5p suppresses PDZK1IP1 expression and mediates OSCC progression. MiR-455-5p and PDZK1IP1 may therefore serve as key biomarkers and be involved in regulating partial EMT in OSCC cells. PDZK1IP1 expression may also serve as an independent factor that impacts outcomes in patients with clinical risk factors for recurrence.