Integrated analysis of lncRNA, miRNA and mRNA expression profiles reveals regulatory pathways associated with pig testis function.

Long noncoding RNA (lncRNA) and microRNA (miRNA) are known to play pivotal roles in mammalian testicular function and spermatogenesis. However, their impact on porcine male reproduction has yet to be well unraveled. Here, we sequenced and identified lncRNA and miRNA expressed in the testes of Chinese indigenous Banna mini-pig inbred line (BMI) and introduced Western Duroc (DU) and Large White (LW) pigs. By pairwise comparison (BMI vs DU, BMI vs LW, and DU vs LW), we found the gene expression differences in the testes between Chinese local pigs and introduced Western commercial breeds were more striking than those between introduced commercial breeds. Furthermore, we found 1622 co-differentially expressed genes (co-DEGs), 122 co-differentially expressed lncRNAs (co-DELs), 39 co-differentially expressed miRNAs (co-DEMs) in BMI vs introduced commercial breeds (DU and LW). Functional analysis revealed that these co-DEGs and co-DELs/co-DEMs target genes were enriched in male sexual function pathways, including MAPK, AMPK, TGF-ß/Smad, Hippo, NF-kappa B, and PI3K/Akt signaling pathways. Additionally, we established 10,536 lncRNA-mRNA, 11,248 miRNA-mRNA pairs, and 62 ceRNA (lncRNA-miRNA-mRNA) networks. The ssc-miR-1343 had the most interactive factors in the ceRNA network, including 20 mRNAs and 3 lncRNAs, consisting of 56 ceRNA pairs. These factors played extremely important roles in the regulation of testis function as key nodes in the interactive regulatory network. Our results provide insight into the functional roles of lncRNAs and miRNAs in porcine testis and offer a valuable resource for understanding the differences between Chinese indigenous and introduced Western pigs.

Genome-Wide Detection for Runs of Homozygosity in Baoshan Pigs Using Whole Genome Resequencing.

Baoshan pigs (BS) are a local breed in Yunnan Province that may face inbreeding owing to its limited population size. To accurately evaluate the inbreeding level of the BS pig population, we used whole-genome resequencing to identify runs of homozygosity (ROH) regions in BS pigs, calculated the inbreeding coefficient based on pedigree and ROH, and screened candidate genes with important economic traits from ROH islands. A total of 22,633,391 SNPS were obtained from the whole genome of BS pigs, and 201 ROHs were detected from 532,450 SNPS after quality control. The number of medium-length ROH (1-5 Mb) was the highest (98.43%), the number of long ROH (>5 Mb) was the lowest (1.57%), and the inbreeding of BS pigs mainly occurred in distant generations. The inbreeding coefficient FROH, calculated based on ROH, was 0.018 ± 0.016, and the FPED, calculated based on the pedigree, was 0.027 ± 0.028, which were positively correlated. Forty ROH islands were identified, containing 507 genes and 891 QTLs. Several genes were associated with growth and development (IGFALS, PTN, DLX5, DKK1, WNT2), meat quality traits (MC3R, ACSM3, ECI1, CD36, ROCK1, CACNA2D1), and reproductive traits (NPW, TSHR, BMP7). This study provides a reference for the protection and utilization of BS pigs.

Endoscopic thyroidectomy via areola approach for stage T1 papillary thyroid carcinoma: feasibility, safety, and oncologic outcomes.

Purpose: To evaluate the feasibility, safety, and oncologic outcomes associated with endoscopic thyroidectomy via the areolar approach (ETAA), compared with conventional open thyroidectomy (COT) for the treatment of stage T1 papillary thyroid carcinoma (PTC). Methods: Between January 2021 and June 2022, a total of 1204 patients diagnosed with PTC underwent screening, out of which 138 patients were selected for inclusion in the study population after propensity score matching (92 patients in the ETAA group and 46 patients in the COT group). The study included the collection and analysis of clinicopathologic characteristics, intraoperative outcomes, postoperative outcomes, complications, and follow-up data using R software. Results: The operative time for the ETAA group was longer than that for the COT group (160.42 ± 32.21 min vs. 121.93 ± 29.78 min, p < 0.0001). However, there were no significant differences between the two groups in terms of intraoperative blood loss, the extent of surgical resection, the number of dissected lymph nodes, the number of metastatic lymph nodes, and the rate of parathyroid autotransplantation. Postoperative drainage and C-reactive protein levels were higher in the ETAA group than in the COT group, but there were no significant differences in 24-hour visual analogue scale scores, white blood cell counts, drainage duration, or postoperative hospital stay. Complication rates were similar between the two groups, and no permanent recurrent laryngeal nerve palsy or hypoparathyroidism was observed. Patients who underwent ETAA reported greater cosmetic satisfaction and quality of life than those who underwent COT. During the follow-up phase, only one patient in the COT group developed lateral cervical lymph node involvement requiring reoperation. Conclusion: ETAA is a safe and feasible surgical method for patients with stage T1 PTC, providing results similar to COT in terms of oncologic completeness, while avoiding neck scars, with excellent cosmetic effects. Clinical trial registration: Chinese Clinical Trial Registry center, identifier ChiCTR2300077109.

Causal association between body mass index and autoimmune thyroiditis: evidence from Mendelian randomization.

BACKGROUND: Recent studies have reported associations between body mass index (BMI) and various autoimmune disorders. However, it is still uncertain whether there exists a direct cause-and-effect relationship between BMI and autoimmune thyroiditis (AIT). The aim of our study is to investigate the causal association between BMI and AIT. METHODS: We conducted a two-sample summary data Mendelian randomization (MR) analysis using genome-wide association studies (GWAS) summary statistics data related to BMI as exposure, and GWAS summary statistic data sets for AIT as the outcome. Robustly associated single-nucleotide polymorphisms (SNPs) for BMI were selected as instrumental variables (IVs). We used the inverse variance weighted (IVW) method as the primary method and performed other MR methods such as MR-Egger regression, weighted median, simple mode, and weighted mode analyses for further validation. The slope of MR-Egger regression was used to correct for pleiotropy and provide estimates of causality. The p-value for the intercept in MR-Egger was utilized to detect any directional pleiotropic effects. Heterogeneity and sensitivity analyses were performed to assess the robustness of our findings. RESULTS: Seventy-eight SNPs were selected from GWAS on BMI as the IVs. Our MR analysis using the IVW method showed a potential causal association between BMI and AIT (OR = 3.071, 95% CI 1.324-7.118). Findings from other MR methods are non-significant, although the direction of effect is consistent. There was no evidence that the result was affected by genetic pleiotropy (MR-Egger regression intercept = 0.01, SE = 0.00025, p = 0.719). Heterogeneity and sensitivity analyses revealed no significant heterogeneity among SNPs, and no single SNP drove the observed associations. CONCLUSION: Our findings suggest a potential causal association between BMI and AIT, which may provide a basis for further investigation into the relationship between BMI and AIT. Further studies are required as only the IVW method shows significant results, and the case sample size is small.

Transoral Endoscopic Thyroidectomy Vestibular Approach (TOETVA): Influences on the Voice Changes and Swallowing Function Disorders.

OBJECTIVE: The aim of this study was to evaluate the influence of transoral endoscopic thyroidectomy vestibular approach (TOETVA) on voice changes and swallowing function disorders. MATERIALS AND METHODS: We retrospectively reviewed 215 patients who underwent thyroid surgery with TOETVA (105 cases, endoscopic group) and open approach (110 cases, open group). Major outcomes, the changes in voice and swallowing function in the 2 groups of patients before and after surgery were analyzed by using both subjective and objective evaluation indexes. Subjective evaluation indexes included the Voice Handicap Index, voice GRBAS (Grade, Roughness, Breathiness, Asthenia, Strain) classification, and swallowing impairment score; the objective indicators included the fundamental frequency (F0), fundamental frequency perturbation (jitter), amplitude perturbation (shimmer), and maximum phonation time. RESULTS: In terms of subjective evaluation indexes, there were no significant differences (all P> 0.05) between the groups regarding Voice Handicap Index (1 and 3 mo after surgery) and GRBAS (1 and 3 mo after surgery). The incidence rates of swallowing disorder in the endoscopic group were higher than that in the open group according to the outcomes of swallowing impairment score at 1 and 3 months after surgery (both P< 0.05). In addition, no significant changes in terms of jitter, shimmer, and maximum phonation time in both groups of patients at 1 and 3 months after surgery compared with their preoperative values (all P> 0.05). CONCLUSIONS: Voice and swallowing disorders may occur in some patients, either TOETVA or open thyroid surgery, which in most cases will recover within 3 months after surgery. The time to swallowing function recovery is relatively prolonged in patients following TOETVA, which may be probably associated with neck adhesion and fixation after the operation.

Long- and short-read RNA sequencing from five reproductive organs of boar.

The production of semen in boars involves multiple reproductive glands, including the testis (Tes), epididymis (Epi), vesicular gland (VG), prostate gland (PG), and bulbourethral gland (BG). However, previous studies on boar reproduction primarily focused on the testis, with little attention paid to the other glands. Here, we integrated single-molecule long-read sequencing with short-read sequencing to characterize the RNA landscape from five glands of Banna mini-pig inbred line (BMI) and Diannan small-ear pigs (DSE). We identified 110,996 full-length isoforms from 22,298 genes, and classified the alternative splicing (AS) events in these five glands. Transcriptome-wide variation analysis indicated that the number of single nucleotide polymorphisms (SNPs) in five tissues of BMI was significantly lower than that in the non-inbred pig, DSE, revealing the effect of inbreeding on BMI. Additionally, we performed small-RNA sequencing and identified 299 novel miRNAs across all glands. Overall, our findings provide a comprehensive overview of the RNA landscape within these five glands, paving the path for future investigations on reproductive biology and the impact of inbreeding on pig transcriptome.

Safety and Efficacy of Transoral Endoscopic Thyroidectomy Vestibular Approach Versus Conventional Open Thyroidectomy: A Systematic Review and Meta-analysis.

OBJECTIVE: To evaluate the safety and efficacy of the transoral endoscopic thyroidectomy vestibular approach (TOETVA) versus conventional open thyroidectomy (COT) for some thyroid diseases. MATERIALS AND METHODS: Databases PubMed, Embase, and Web of Science were searched. Full-text English papers that described TOETVA and COT for people with thyroid diseases were included. Randomized, nonrandomized, controlled, and uncontrolled trials were all included. Extracted data included population characteristics and intraoperative and postoperative complications. RESULTS: A total of 2 randomized controlled trials and 10 retrospective studies, including 3048 patients, were included in the meta-analysis. Meta-analysis results suggested that the intraoperative conditions and postoperative complication rates did not differ significantly between the two groups. However, in the TOETVA group, there is a slightly longer operative time [weighted mean difference (WMD): 73.64; 95% CI: 49.34 to 97.94; P < 0.0001], drainage (WMD: 91.0; 95% CI: 35.52 to 146.48; P = 0.001), and hospital stay (WMD: 0.28; 95% CI: 0.18 to 0.38; P < 0.0001). CONCLUSION: For most of the benign thyroid nodules and selected patients with papillary thyroid cancer, TOETVA seems to be as feasible and safe as COT.

Effects of Different Levels of Surgical Separation on Neck Bulge and Swallowing Disorders during Endoscopic Thyroidectomy.

OBJECTIVE: Our study aims to compare the effects of different levels of access on postoperative neck bulge and swallowing disorder and provide a better level for endoscopic thyroidectomy. MATERIALS AND METHODS: The patients were selected retrospectively between March 2021 to September 2021 by the Department of Thyroid Surgery, Third Affiliated Hospital of Zunyi Medical University. They were divided into two groups according to the level of the free flap during surgery: group A (the superficial cervical fascial level) and group B (the superficial deep cervical fascial level). Age, sex, body mass index, diameter of the primary lesion, postoperative neck bulge, and swallowing disorders and other complications were compared between the two groups. RESULTS: A total of 40 patients who underwent endoscopic unilateral lobectomy plus central region lymph node dissection were enrolled in our study. Twenty in group A and 20 in group B. The age, gender, body mass index, diameter, and the proportion of benign and malignant primary lesions were not significantly different between the two groups ( P >0.05), and there was also no difference in thyroid function between the two groups ( P >0.05). No significant differences were observed in terms of bleeding and operation time during the operation ( P >0.05). There were also no statistical differences in terms of recurrent laryngeal nerve injury or hypoparathyroidism ( P >0.05). However, patients in group B were superior to those in group A in the occurrence of neck bulge and swallowing disorders ( P <0.05). These symptoms were most evident one month after surgery. Six months after the operation, only 4 patients in group B still complained of neck "swelling" and uncomfortable straining which did not resolve until 1 year after the operation. No statistical significance between long-term results and complication rates could be observed in either group. CONCLUSION: In endoscopic thyroidectomy, the superficial cervical fascial level may be a better option for reducing postoperative neck bulge and swallowing disorders, which needs further validation by conducting a large sample study.

Molecular characteristics and transcriptional regulatory of spermatogenesis-related gene RFX2 in adult Banna mini-pig inbred line (BMI).

RFX2 plays critical roles in mammalian spermatogenesis and cilium maturation. Here, the testes of 12-month-old adult boars of Banna mini-pig inbred line (BMI) were subjected to whole-transcriptome sequencing. The results indicated that the average expression (raw count) of RFX2 gene in BMI testes was 16138.25, and the average expression value of the corresponding transcript ENSSSCT00000043271.2 was 123.1898. The CDS of RFX2 obtained from BMI testes was 2,817 bp (GenBank accession number: OL362242). Gene structure analysis showed that RFX2 was located on chromosome 2 of the pig genome with 19 exons. Protein structure analysis indicated that RFX2 contains 728 amino acids with two conserved domains. Phylogenetic analysis revealed that RFX2 was highly conserved with evolutionary homologies among mammalian species. Other analyses, including PPI networks, KEGG, and GO, indicated that BMI RFX2 had interactions with 43 proteins involving various functions, such as in cell cycle, spermatid development, spermatid differentiation, cilium assembly, and cilium organization, etc. Correlation analysis between these proteins and the transcriptome data implied that RFX2 was significantly associated with FOXJ1, DNAH9, TMEM138, E2F7, and ATR, and particularly showed the highest correlation with ATR, demonstrating the importance of RFX2 and ART in spermatogenesis. Functional annotation implied that RFX2 was involved in 17 GO terms, including three cellular components (CC), six molecular functions (MF), and eight biological processes (BP). The analysis of miRNA-gene targeting indicated that BMI RFX2 was mainly regulated by two miRNAs, among which four lncRNAs and five lncRNAs competitively bound ssc-miR-365-5p and ssc-miR-744 with RFX2, respectively. Further, the dual-luciferase report assay indicated that the ssc-miR-365-5p and ssc-miR-744 significantly reduced luciferase activity of RFX2 3'UTR in the 293T cells, suggesting that these two miRNAs regulated the expression of RFX2. Our results revealed the important role of RFX2 in BMI spermatogenesis, making it an intriguing candidate for follow-up studies.

Ultrasound-Guided Capsular Thyroid Injection Therapy With Dexamethasone and Lidocaine Mixture for Subacute Thyroiditis: A Single-Center Study.

OBJECTIVES: Subacute thyroiditis (SAT) is a self-limiting, inflammatory thyroid disease possibly caused by viral infection. In recent years, the incidence of SAT is increasing, especially during the pandemic of the COVID-19. This study aimed to evaluate the efficacy, safety, and recovery time of capsular thyroid injection therapy under ultrasound guidance for SAT. METHODS: A total of 73 patients with SAT were divided into two groups. Patients in group A (n = 48) received an ultrasound-guided capsular injection consisting of dexamethasone (DEX) and lidocaine in the thyroid lesion area, while patients in group B (n = 25) received oral prednisolone (PSL). The two groups were compared for pain relief and treatment duration, the recovery time of thyroid function, recurrence rates, hypothyroidism incidence, and drug-related side effects. RESULTS: The follow-up time was 1 year. In group A, the duration of pain relief, treatment, and recovery time of thyroid function were significantly shorter than that in group B (P < .05), and no statistically significant differences in recurrence rate or incidence of hypothyroidism were observed (P > .05). Weight gain was significantly higher in group A at the end of treatment (P < .001). CONCLUSIONS: Compared with oral PSL treatment, ultrasound-guided local injection of DEX and lidocaine into the capsular thyroid is a safe and effective procedure that can significantly reduce the treatment time of SAT.

Transcriptomic analysis of testis and epididymis tissues from Banna mini-pig inbred line boars with single-molecule long-read sequencing†.

In mammals, testis and epididymis are critical components of the male reproductive system for androgen production, spermatogenesis, sperm transportation, as well as sperm maturation. Here, we report single-molecule real-time sequencing data from the testis and epididymis of the Banna mini-pig inbred line (BMI), a promising laboratory animal for medical research. We obtained high-quality full-length transcriptomes and identified 9879 isoforms and 8761 isoforms in the BMI testis and epididymis, respectively. Most of the isoforms we identified have novel exon structures that will greatly improve the annotation of testis- and epididymis-expressed genes in pigs. We also found that 3055 genes (over 50%) were shared between BMI testis and epididymis, indicating widespread expression profiles of genes related to reproduction. We characterized extensive alternative splicing events in BMI testis and epididymis and showed that 96 testis-expressed genes and 79 epididymis-expressed genes have more than six isoforms, revealing the complexity of alternative splicing. We accurately defined the transcribed isoforms in BMI testis and epididymis by combining Pacific Biotechnology Isoform-sequencing (PacBio Iso-Seq) and Illumina RNA Sequencing (RNA-seq) techniques. The refined annotation of some key genes governing male reproduction will facilitate further understanding of the molecular mechanisms underlying BMI male sterility. In addition, the high-confident identification of 548 and 669 long noncoding RNAs (lncRNAs) in these two tissues has established a candidate gene set for future functional investigations. Overall, our study provides new insights into the role of the testis and epididymis during BMI reproduction, paving the path for further studies on BMI male infertility.

Inverse association between type 2 diabetes and hepatocellular carcinoma in East Asian populations.

Aims: To investigate the potential association between type 2 diabetes (T2D) and hepatocellular carcinoma (HCC) in East Asian populations using Mendelian randomization (MR) analyses. Methods: Bidirectional Mendelian randomization (MR) studies were conducted using summary statistics from genome-wide association studies (GWAS) related to T2D and HCC. The potential effects of confounders such as chronic hepatitis B, chronic hepatitis C, body mass index, and alcohol intake frequency were corrected using a multivariate MR study. Various MR methods, including the inverse variance weighted (IVW) approach, were used to estimate the associations between T2D and HCC. Sensitivity analysis and assessment of heterogeneity were performed to ensure the robustness of the results. Results: In the forward MR study, the IVW approach of MR analysis suggested an inverse association between T2D and HCC, with a risk odds ratio of 0.8628 (95% confidence interval [CI], 0.7888-0.9438). Furthermore, even after adjusting for BMI, chronic hepatitis B, and alcohol intake frequency, this study still supports the inverse association between T2D and HCC. Additional MR methods provided further support for this relationship. Sensitivity analysis and assessment of heterogeneity confirmed the robustness of the results. The reverse MR analysis did not show a clear impact of genetic liability to HCC on reduced risk of T2D(OR=0.9788; 95% CI, 0.9061-1.0574). Conclusion: This study provides evidence of an inverse association between T2D and HCC in East Asian populations using MR analysis. Further studies are warranted to validate these findings.

Factors Influencing the Acceptance of Endoscopic Thyroidectomy Via Oral Vestibular Approach (ETOVA) by Thyroid Surgery Candidates.

OBJECTIVE: To investigate the factors affecting the acceptance of endoscopic thyroidectomy via the oral vestibular approach (ETOVA) in Chinese patients before thyroid surgery. METHODS: The enrolled patients were asked to answer a questionnaire postoperatively about their demographics, medical insurance coverage, sources of information, reasons for selection, and safety. The relationship between the collected data and the acceptance of ETOVA was analyzed. RESULTS: Two hundred patients (40 males, 20%; 160 females, 80%) answered the questionnaire. One hundred sixty-two of them (81%) accepted ETOVA. Univariate analysis showed that the patients' age, cosmetic effect, safety, results perception, and recommendations from family, friends, doctors, and nurses are correlated with the acceptance of ETOVA. Multivariate analysis showed that patients' age (OR=0.966, P =0.015), cosmetic effect (OR=12.620, P =0.000), safety (OR=0.295, P =0.016), minimal invasion (OR=4.877, P =0.001), and doctors/nurses' advance (OR=4.485, P =0.017) are statistically significant and were positively correlated with the acceptance of ETOVA. Education level, medical insurance coverage, family support, past surgical history, and operative-related symptoms were not statistically significant ( P >0.05). CONCLUSION: Among thyroid surgery candidates in Southwest China, younger patients with cosmetic requirements and minimally invasive procedures desires are more likely to consider ETOVA at the urging of their physicians/nurses. Providing appropriate healthcare education, medical insurance coverage, and information options for surgical treatments is vital to improving patients' acceptance of ETOVA.

Dynamic 3D genome reorganization during development and metabolic stress of the porcine liver.

Liver development is a complex process that is regulated by a series of signaling pathways. Three-dimensional (3D) chromatin architecture plays an important role in transcriptional regulation; nonetheless, its dynamics and role in the rapid transition of core liver functions during development and obesity-induced metabolic stress remain largely unexplored. To investigate the dynamic chromatin architecture during liver development and under metabolic stress, we generated high-resolution maps of chromatin architecture for porcine livers across six major developmental stages (from embryonic day 38 to the adult stage) and under a high-fat diet-induced obesity. The characteristically loose chromatin architecture supports a highly plastic genome organization during early liver development, which fundamentally contributes to the rapid functional transitions in the liver after birth. We reveal the multi-scale reorganization of chromatin architecture and its influence on transcriptional regulation of critical signaling processes during liver development, and show its close association with transition in hepatic functions (i.e., from hematopoiesis in the fetus to metabolism and immunity after birth). The limited changes in chromatin structure help explain the observed metabolic adaptation to excessive energy intake in pigs. These results provide a global overview of chromatin architecture dynamics associated with the transition of physiological liver functions between prenatal development and postnatal maturation, and a foundational resource that allows for future in-depth functional characterization.

Ethylene-Inducible AP2/ERF Transcription Factor Involved in the Capsaicinoid Biosynthesis in Capsicum.

Ethylene is very important in the process of plant development and regulates the biosynthesis of many secondary metabolites. In these regulatory mechanisms, transcription factors (TFs) that mediate ethylene signals play a very important role. Capsaicinoids (CAPs) are only synthesized and accumulated in Capsicum species, causing their fruit to have a special pungent taste, which can protect against attack from herbivores and pathogens. In this study, we identified the TF CcERF2, which is induced by ethylene, and demonstrated its regulatory effect on CAPs biosynthesis. Transcriptome sequencing analysis revealed that the expression patterns of CcERF2 and multiple genes associated with CAPs biosynthesis were basically the same. The spatiotemporal expression results showed CcERF2 was preferentially expressed in the placenta of the spicy fruit. Ethylene can induce the expression of CcERF2 and CAPs biosynthesis genes (CBGs). CcERF2 gene silencing and 1-methylcyclopropene (1-MCP) and pyrazinamide (PZA) treatments caused a decrease in expression of CBGs and a sharp decrease in content of CAPs. The results indicated that CcERF2 was indeed involved in the regulation of structural genes of the CAPs biosynthetic pathway.