CDK11 requires a critical activator SAP30BP to regulate pre-mRNA splicing.

CDK11 is an emerging druggable target for cancer therapy due to its prevalent roles in phosphorylating critical transcription and splicing factors and in facilitating cell cycle progression in cancer cells. Like other cyclin-dependent kinases, CDK11 requires its cognate cyclin, cyclin L1 or cyclin L2, for activation. However, little is known about how CDK11 activities might be modulated by other regulators. In this study, we show that CDK11 forms a tight complex with cyclins L1/L2 and SAP30BP, the latter of which is a poorly characterized factor. Acute degradation of SAP30BP mirrors that of CDK11 in causing widespread and strong defects in pre-mRNA splicing. Furthermore, we demonstrate that SAP30BP facilitates CDK11 kinase activities in vitro and in vivo, through ensuring the stabilities and the assembly of cyclins L1/L2 with CDK11. Together, these findings uncover SAP30BP as a critical CDK11 activator that regulates global pre-mRNA splicing.

The polyA tail facilitates splicing of last introns with weak 3' splice sites via PABPN1.

The polyA tail of mRNAs is important for many aspects of RNA metabolism. However, whether and how it regulates pre-mRNA splicing is still unknown. Here, we report that the polyA tail acts as a splicing enhancer for the last intron via the nuclear polyA binding protein PABPN1 in HeLa cells. PABPN1-depletion induces the retention of a group of introns with a weaker 3' splice site, and they show a strong 3'-end bias and mainly locate in nuclear speckles. The polyA tail is essential for PABPN1-enhanced last intron splicing and functions in a length-dependent manner. Tethering PABPN1 to nonpolyadenylated transcripts also promotes splicing, suggesting a direct role for PABPN1 in splicing regulation. Using TurboID-MS, we construct the PABPN1 interactome, including many spliceosomal and RNA-binding proteins. Specifically, PABPN1 can recruit RBM26&27 to promote splicing by interacting with the coiled-coil and RRM domain of RBM27. PABPN1-regulated terminal intron splicing is conserved in mice. Together, our study establishes a novel mode of post-transcriptional splicing regulation via the polyA tail and PABPN1.

Highly multiplex PCR assays by coupling the 5'-flap endonuclease activity of Taq DNA polymerase and molecular beacon reporters.

Real-time PCR is the most utilized nucleic acid testing tool in clinical settings. However, the number of targets detectable per reaction are restricted by current modes. Here, we describe a single-step, multiplex approach capable of detecting dozens of targets per reaction in a real-time PCR thermal cycler. The approach, termed MeltArray, utilizes the 5'-flap endonuclease activity of Taq DNA polymerase to cleave a mediator probe into a mediator primer that can bind to a molecular beacon reporter, which allows for the extension of multiple mediator primers to produce a series of fluorescent hybrids of different melting temperatures unique to each target. Using multiple molecular beacon reporters labeled with different fluorophores, the overall number of targets is equal to the number of the reporters multiplied by that of mediator primers per reporter. The use of MeltArray was explored in various scenarios, including in a 20-plex assay that detects human Y chromosome microdeletions, a 62-plex assay that determines Escherichia coli serovars, a 24-plex assay that simultaneously identifies and quantitates respiratory pathogens, and a minisequencing assay that identifies KRAS mutations, and all of these different assays were validated with clinical samples. MeltArray approach should find widespread use in clinical settings owing to its combined merits of multiplicity, versatility, simplicity, and accessibility.

Intrinsically disordered interaction network in an RNA chaperone revealed by native mass spectrometry.

RNA-binding proteins contain intrinsically disordered regions whose functions in RNA recognition are poorly understood. The RNA chaperone Hfq is a homohexamer that contains six flexible C-terminal domains (CTDs). The effect of the CTDs on Hfq's integrity and RNA binding has been challenging to study because of their sequence identity and inherent disorder. We used native mass spectrometry coupled with surface-induced dissociation and molecular dynamics simulations to disentangle the arrangement of the CTDs and their impact on the stability of Escherichia coli Hfq with and without RNA. The results show that the CTDs stabilize the Hfq hexamer through multiple interactions with the core and between CTDs. RNA binding perturbs this network of CTD interactions, destabilizing the Hfq ring. This destabilization is partially compensated by binding of RNAs that contact multiple surfaces of Hfq. By contrast, binding of short RNAs that only contact one or two subunits results in net destabilization of the complex. Together, the results show that a network of intrinsically disordered interactions integrate RNA contacts with the six subunits of Hfq. We propose that this CTD network raises the selectivity of RNA binding.

Plasmodium yoelii surface-related antigen (PySRA) modulates the host pro-inflammatory responses via binding to CD68 on macrophage membrane.

Malaria, one of the major infectious diseases in the world, is caused by the Plasmodium parasite. Plasmodium antigens could modulate the inflammatory response by binding to macrophage membrane receptors. As an export protein on the infected erythrocyte membrane, Plasmodium surface-related antigen (SRA) participates in the erythrocyte invasion and regulates the immune response of the host. This study found that the F2 segment of P. yoelii SRA activated downstream MAPK and NF-κB signaling pathways by binding to CD68 on the surface of the macrophage membrane and regulating the inflammatory response. The anti-PySRA-F2 antibody can protect mice against P. yoelii, and the pro-inflammatory responses such as IL-1ß, TNF-α, and IL-6 after infection with P. yoelii are attenuated. These findings will be helpful for understanding the involvement of the pathogenic mechanism of malaria with the exported protein SRA.

Protein analysis using capillary electrophoresis coupled to mass spectrometry through vibrating sharp-edge spray ionization.

Capillary electrophoresis (CE) interfaced to mass spectrometry (MS) with electrospray ionization typically incorporates acidic additives or organic solvents to assist in ionization. Vibrating sharp-edge spray ionization (VSSI) is a voltage-free method to interface CE and MS that does not require these additives, making it appealing for protein analyses. CE-VSSI nanoflow sheath separations are performed with low ionic strength aqueous solutions in the sheath to reduce suppression. Serine is also included in the sheath to reduce analyte adduction. Proteins are detected in the 2.5-10 µM range, corresponding to an injected mass range of 0.1-1.2 ng. The anionic proteins ß-lactoglobulin and transferrin are resolved using an unmodified fused silica capillary because they do not exhibit nonspecific surface adsorption. Conversely, separations of cationic proteins cytochrome c, ribonuclease A, and α-chymotrypsinogen A in an unmodified capillary require acidic background electrolytes to overcome adsorption. Alternatively, a semipermanent coating comprised self-assembled lipids overcomes surface adsorption at a neutral pH. Separations with zwitterionic and hybrid cationic coatings are complete within 15 or 6 min, respectively. The dimeric form of triosephosphate isomerase was observed at a 60 µM, corresponding to a mass of 19 ng, by dropping the temperature of the MS inlet.

Endoscopic resection for the treatment of gastric gastrointestinal stromal tumors: a retrospective study from a large tertiary hospital in China.

BACKGROUND AND AIMS: With the continuous development of endoscopic technology, endoscopic resection (ER) has gradually become an optional method for the treatment of gastric gastrointestinal stromal tumors (GISTs). However, studies with a large sample or a long follow-up are lacking. Therefore, this research aims to evaluate the efficacy and safety of ER for gastric GISTs in the real-world setting with more than 300 enrolled patients and a follow-up period longer than 45 months. METHODS: From January 2013 to February 2023, 409 patients with a pathological diagnosis of GISTs after ER were retrospectively enrolled in this study. After excluding 86 patients with non-gastric GISTs, we assessed 323 patients with gastric GISTs. The main outcome measures were en bloc resection, complete resection, residual disease, recurrence, and complications. RESULTS: There were 194 (60.06%) females and 129 (39.94%) males, and the median age of the included patients was 58 years (51, 63). The median tumor size was 15.0 (10.0, 20.0) mm. According to the modified NIH criteria, 246 (75.85%) patients were classified as very low risk, 62 (19.20%) were classified as low risk, 12 (3.72%) were classified as moderate risk, and 3 (0.93%) were classified as high risk. A total of 287 (88.85%) patients achieved en bloc resection, and 287 (88.85%) also achieved complete resection. Only one patient showed residual and no recurrent lesions were noted during the follow-up. Regarding complications, three patients had complications, with a complication rate of 0.93%, and no severe complications requiring surgical intervention occurred. CONCLUSION: ER is an appropriate alternative method for the treatment of gastric GISTs, with an en bloc resection rate of 88.85% and a complication rate of 0.93%. No recurrence was noted during follow-up, even for GISTs with piecemeal resection.

Elucidation of structure-function relationships in Methanocaldococcus jannaschii RNase P, a multi-subunit catalytic ribonucleoprotein.

RNase P is a ribonucleoprotein (RNP) that catalyzes removal of the 5' leader from precursor tRNAs in all domains of life. A recent cryo-EM study of Methanocaldococcus jannaschii (Mja) RNase P produced a model at 4.6-Å resolution in a dimeric configuration, with each holoenzyme monomer containing one RNase P RNA (RPR) and one copy each of five RNase P proteins (RPPs; POP5, RPP30, RPP21, RPP29, L7Ae). Here, we used native mass spectrometry (MS), mass photometry (MP), and biochemical experiments that (i) validate the oligomeric state of the Mja RNase P holoenzyme in vitro, (ii) find a different stoichiometry for each holoenzyme monomer with up to two copies of L7Ae, and (iii) assess whether both L7Ae copies are necessary for optimal cleavage activity. By mutating all kink-turns in the RPR, we made the discovery that abolishing the canonical L7Ae-RPR interactions was not detrimental for RNase P assembly and function due to the redundancy provided by protein-protein interactions between L7Ae and other RPPs. Our results provide new insights into the architecture and evolution of RNase P, and highlight the utility of native MS and MP in integrated structural biology approaches that seek to augment the information obtained from low/medium-resolution cryo-EM models.

Bi-Allelic UQCRFS1 Variants Are Associated with Mitochondrial Complex III Deficiency, Cardiomyopathy, and Alopecia Totalis.

Isolated complex III (CIII) deficiencies are among the least frequently diagnosed mitochondrial disorders. Clinical symptoms range from isolated myopathy to severe multi-systemic disorders with early death and disability. To date, we know of pathogenic variants in genes encoding five out of 10 subunits and five out of 13 assembly factors of CIII. Here we describe rare bi-allelic variants in the gene of a catalytic subunit of CIII, UQCRFS1, which encodes the Rieske iron-sulfur protein, in two unrelated individuals. Affected children presented with low CIII activity in fibroblasts, lactic acidosis, fetal bradycardia, hypertrophic cardiomyopathy, and alopecia totalis. Studies in proband-derived fibroblasts showed a deleterious effect of the variants on UQCRFS1 protein abundance, mitochondrial import, CIII assembly, and cellular respiration. Complementation studies via lentiviral transduction and overexpression of wild-type UQCRFS1 restored mitochondrial function and rescued the cellular phenotype, confirming UQCRFS1 variants as causative for CIII deficiency. We demonstrate that mutations in UQCRFS1 can cause mitochondrial disease, and our results thereby expand the clinical and mutational spectrum of CIII deficiencies.

Combining Surface-Induced Dissociation and Charge Detection Mass Spectrometry to Reveal the Native Topology of Heterogeneous Protein Complexes.

Charge detection mass spectrometry (CDMS) enables the direct mass measurement of heterogeneous samples on the megadalton scale, as the charge state for a single ion is determined simultaneously with the mass-to-charge ratio (m/z). Surface-induced dissociation (SID) is an effective activation method to dissociate non-intertwined, non-covalent protein complexes without extensive gas-phase restructuring, producing various subcomplexes reflective of the native protein topology. Here, we demonstrate that using CDMS after SID on an Orbitrap platform offers subunit connectivity, topology, proteoform information, and relative interfacial strengths of the intact macromolecular assemblies. SID dissects the capsids (∼3.7 MDa) of adeno-associated viruses (AAVs) into trimer-containing fragments (3mer, 6mer, 9mer, 15mer, etc.) that can be detected by the individual ion mass spectrometry (I2MS) approach on Orbitrap instruments. SID coupled to CDMS provides unique structural insights into heterogeneous assemblies that are not readily obtained by traditional MS measurements.

Identification and expression analysis of invertase family genes during grape (Vitis vinifera L.) berry development under CPPU and GA treatment.

Sugar is crucial for grape berry, whether used for fresh food or wine. However, berry enlargement treatment with forchlorfenuron (N-(2-chloro4-pyridyl)-N'-phenylurea) (CPPU, a synthetic cytokinin) and gibberellin (GA) always had adverse effects on sugar accumulation in some grape varieties, especially CPPU. Therefore exploring the molecular mechanisms behind these adverse effects could provide a foundation for improving or developing technology to mitigate the effects of CPPU/GA treatments for grape growers. In the present study, invertase (INV) family, the key gene controlling sugar accumulation, was identified and characterized on the latest annotated grape genome. Their express pattern, as well as invertase activity and sugar content, were analyzed during grape berry development under CPPU and GA3 treatment to explore the potential role of INV members under berry enlargement treatment in grapes. Eighteen INV genes were identified and divided into two sub-families: 10 neutral INV genes (Vv-A/N-INV1-10) and 8 acid INV genes containing 5 CWINV (VvCWINV1-5) and 3 VIN (VvVIN1-3). At the early development stage, both CPPU and GA3 treatment decreased the hexose level in berries of 'Pinot Noir' grape, whereas the activity of three types inverstase (soluble acid INV, insoluble acid INV, and neutral INV) increased. Correspondingly, most of INV members were up-regulated by GA3 /CPPU application at least one sampling time point during early berry development, including VvCWINV1, 2, 3, 4, 5, VvVIN1, 2, 3 and Vv-A/N-INV1, 2, 5, 6, 7, 8, 10. At maturity, the sugar content in CPPU-treated berries is still lower than that in the control. Soluble acid INV and neutral INV, rather than insoluble acid INV, presented lower activity in CPPU-treated berries. Meanwhile, several corresponding genes, such as VvVIN2 and Vv-A/N-INV2, 8, 10 in ripening berries were obviously down-regulated by CPPU treatment. These results suggested that most of INV members could be triggered by berry enlargement treatment during early berry development, whereas VvVINs and Vv-A/N-INVs, but not VvCWINVs, could be the limiting factor resulting in decreased sugar accumulation in CPPU-treated berries at maturity. In conclusion, this study identified the INV family on the latest annotated grape genome and selected several potential members involving in the limit of CPPU on final sugar accumulation in grape berry. These results provide candidate genes for further study of the molecular regulation of CPPU and GA on sugar accumulation in grape.

Causes of endoscopic misdiagnosis of gastrointestinal cyst as solid lesion.

BACKGROUND: To explore the causes of endoscopic misdiagnosis of gastrointestinal cyst as solid lesion and the diagnostic value and limitations of EUS, guide clinicians to develop appropriate treatment strategies and improve the ability to identify SMT. METHODS: We enrolled patients diagnosed with gastrointestinal SMT between January 2001 and December 2021 who underwent endoscopic resection with postoperative pathological diagnosis of cyst. Age, sex, maximum lesion diameter, judge the texture of lesion, origin and echo are potential factors affecting the diagnostic accuracy of cysts. RESULTS: The diagnostic accuracy of EUS assessment 39.3% higher than that without EUS assessment (6.7%). The error rate was 60.7%, lower than that without EUS assessment (93.3%), suggesting that preoperative EUS assessment improved the diagnostic accuracy of gastrointestinal cyst (Fisher's accurate test, P = 0.033). The diagnostic accuracy of "judge the texture of lesion" was higher than that of no touch (P = 0.031). When the lesion size increased by 1 cm, the diagnostic accuracy decreased by about 21%. Hypoechoic lesions were less likely to be diagnosed correctly than anechoic lesions (P = 0.003). CONCLUSIONS: The main cause of misdiagnosing gastrointestinal cyst as solid lesion is that no EUS assessment was performed before endoscopic resection or anechoic lesion was judged as hypoechoic lesion by preoperative EUS assessment.

Self-help inflatable balloon versus autologous skin-grafting surgery for preventing esophageal stricture after complete circular endoscopic submucosal dissection: a propensity score matching analysis.

BACKGROUND: The self-help inflatable balloon (SHIB) and autologous skin-grafting surgery (ASGS) were used to prevent stricture after esophageal complete circular endoscopic submucosal dissection (cESD) with promising clinical results. We aim to evaluate which method is more suitable for patients who underwent esophageal cESD. METHODS: From October 2017 to July 2021, patients whose mucosal defect length were between 30 and 100 mm after esophageal cESD were retrospectively reviewed from two prospective studies. They were enrolled once SHIB or ASGS was used as preventive methods to prevent stricture. Propensity score matching (PSM) was used to balance the baseline characteristics between the two groups. Comparisons were made between the two groups, including operation time, the longitudinal length of ulceration, fasting time, hospitalization days, and the incidence of stricture. RESULTS: A total of 41 patients who met the inclusion criteria were enrolled in the study. The numbers of patients in SHIB group and ASGS group were 25 and 16, respectively. Fifteen patients in each group were selected after performing PSM. The basic baseline characteristics were comparable between the two groups. The stricture rates were 20% (3/15) in SHIB group and 40% (6/15) in ASGS group, while the difference was not statistically significant (p = 0.427). The SHIB group showed significantly shorter operation time, shorter hospitalization days, lower cost, and longer removing balloon/stent time compared with ASGS group (p < 0.001). Comparison of relevant stricture factors between the stricture group and non-stricture group revealed that longer longitudinal length of ulceration (> 60 mm) accounted for a higher proportion in stricture groups (p = 0.035). CONCLUSION: Both the SHIB and ASGS had high efficacy and safety in preventing strictures in patients with mucosal defects no longer than 100 mm in length after esophageal cESD. The longitudinal length of ulceration > 60 mm was the independent factor for predicting stricture.

Interaction of air pollution and meteorological factors on IVF outcomes: A multicenter study in China.

BACKGROUND: Previous studies revealed associations between air-pollutant exposure and in vitro fertilization (IVF) outcomes. However, modification effects of air pollution on IVF outcomes by meteorological conditions remain elusive. METHODS: This multicenter retrospective cohort study included 15,217 women from five northern Chinese cities during 2015-2020. Daily average concentrations of air pollutants (PM2.5, PM10, O3, NO2, SO2, and CO) and meteorological factors (temperature, relative humidity, wind speed, and sunshine duration) during different exposure windows were calculated as individual approximate exposure. Generalized estimating equations models and stratified analyses were conducted to assess the associations of air pollution and meteorological conditions with IVF outcomes and estimate potential interactions. RESULTS: Positive associations of wind speed and sunshine duration with pregnancy outcomes were detected. In addition, we observed that embryo transfer in spring and summer had a higher likelihood to achieve a live birth compared with winter. Exposure to PM2.5, SO2, and O3 was adversely correlated with pregnancy outcomes in fresh IVF cycles, and the associations were modified by air temperature, relative humidity, and wind speed. The inverse associations of PM2.5 and SO2 exposure with biochemical pregnancy were stronger at lower temperatures and humidity. Negative associations of PM2.5 with clinical pregnancy were only significant at lower temperatures and wind speeds. Moreover, the effects of O3 on live birth were enhanced by higher wind speed. CONCLUSIONS: Our results suggested that the associations between air-pollutant exposure and IVF outcomes were modified by meteorological conditions, especially temperature and wind speed. Women undergoing IVF treatment should be advised to reduce outdoor time when the air quality was poor, particularly at lower temperatures.

The Effect of Prohibitins on Mitochondrial Function during Octopus tankahkeei Spermiogenesis.

Mitochondria are essential for spermiogenesis. Prohibitins (PHBs; prohibitin 1, PHB1 or PHB, and prohibitin 2, PHB2) are evolutionarily conserved and ubiquitously expressed mitochondrial proteins that act as scaffolds in the inner mitochondrial membrane. In this study, we analyzed the molecular structure and dynamic expression characteristics of Ot-PHBs, observed the colocalization of Ot-PHB1 with mitochondria and polyubiquitin, and studied the effect of phb1 knockdown on mitochondrial DNA (mtDNA) content, reactive oxygen species (ROS) levels, and apoptosis-related gene expression in spermatids. Our aim was to explore the effect of Ot-PHBs on mitochondrial function during the spermiogenesis of Octopus tankahkeei (O. tankahkeei), an economically important species in China. The predicted Ot-PHB1/PHB2 proteins contained an N-terminal transmembrane, a stomatin/prohibitin/flotillin/HflK/C (SPFH) domain (also known as the prohibitin domain), and a C-terminal coiled-coil domain. Ot-phb1/phb2 mRNA were widely expressed in the different tissues, with elevated expression in the testis. Further, Ot-PHB1 and Ot-PHB2 were highly colocalized, suggesting that they may function primarily as an Ot-PHB compiex in O. tankahkeei. Ot-PHB1 proteins were mainly expressed and localized in mitochondria during spermiogenesis, implying that their function may be localized to the mitochondria. In addition, Ot-PHB1 was colocalized with polyubiquitin during spermiogenesis, suggesting that it may be a polyubiquitin substrate that regulates mitochondrial ubiquitination during spermiogenesis to ensure mitochondrial quality. To further investigate the effect of Ot-PHBs on mitochondrial function, we knocked down Ot-phb1 and observed a decrease in mtDNA content, along with increases in ROS levels and the expressions of mitochondria-induced apoptosis-related genes bax, bcl2, and caspase-3 mRNA. These findings indicate that PHBs might influence mitochondrial function by maintaining mtDNA content and stabilizing ROS levels; in addition, PHBs might affect spermatocyte survival by regulating mitochondria-induced apoptosis during spermiogenesis in O. tankahkeei.

Expression Dynamics Indicate Potential Roles of KIF17 for Nuclear Reshaping and Tail Formation during Spermiogenesis in Phascolosoma esculenta.

Kinesin family member17 (KIF17), a homologous dimer of the kinesin-2 protein family, has important microtubule-dependent and -independent roles in spermiogenesis. Little is known about KIF17 in the mollusk, Phascolosoma esculenta, a newly developed mariculture species in China. Here, we cloned the open reading frame of Pe-kif17 and its related gene, Pe-act, and performed bioinformatics analysis on both. Pe-KIF17 and Pe-ACT are structurally conserved, indicating that they may be functionally conserved. The expression pattern of kif17/act mRNA performed during spermiogenesis revealed their expression in diverse tissues, with the highest expression level in the coelomic fluid of P. esculenta. The expressions of Pe-kif17 and Pe-act mRNA were relatively high during the breeding season (July-September), suggesting that Pe-KIF17/ACT may be involved in spermatogenesis, particularly during spermiogenesis. Further analysis of Pe-kif17 mRNA via fluorescence in situ hybridization revealed the continuous expression of this mRNA during spermiogenesis, suggesting potential functions in this process. Immunofluorescence showed that Pe-KIF17 co-localized with α-tubulin and migrated from the perinuclear cytoplasm to one side of the spermatid, forming the sperm tail. Pe-KIF17 and Pe-ACT also colocalized. KIF17 may participate in spermiogenesis of P. esculenta, particularly in nuclear reshaping and tail formation by interacting with microtubule structures similar to the manchette. Moreover, Pe-KIF17 with Pe-ACT is also involved in nuclear reshaping and tail formation in the absence of microtubules. This study provides evidence for the role of KIF17 during spermiogenesis and provides theoretical data for studies of the reproductive biology of P. esculenta. These findings are important for spermatogenesis in mollusks.

Diagnostic value of SpyGlass for pancreatic cystic lesions: comparison of EUS-guided fine-needle aspiration and EUS-guided fine-needle aspiration combined with SpyGlass.

BACKGROUND AND AIMS: No study has evaluated the diagnostic value of SpyGlass by comparing SpyGlass results and non-SpyGlass results. In this retrospective study, we aimed to compare the diagnostic value of EUS-guided fine-needle aspiration (EUS-FNA) and EUS-FNA combined with SpyGlass to evaluate whether SpyGlass is valuable for increasing the diagnostic yield of EUS-FNA. METHODS: From April 2015 to April 2020, 251 patients suspected of having pancreatic cystic lesions (PCLs) by imaging techniques who then underwent EUS-FNA were retrospectively enrolled. Only 98 patients who underwent surgical resection with a pathological diagnosis of pancreatic cystic lesion (PCL) were studied. The diagnostic performance outcomes were compared between the EUS-FNA group (EUS-FNA alone, n = 40) and the SpyGlass group (EUS-FNA combined with SpyGlass, n = 58) to assess the value of SpyGlass in diagnosing PCLs. RESULTS: There were 71 females and 27 males with an overall mean age of 47.6 years. The median diameter of the PCLs was 42.2 mm (range, 11.4-100.0 mm). Approximately 37 cysts were localized in the head/neck of the pancreas, while 61 in the body/tail. The sensitivity, specificity, positive predictive value, negative predictive value and diagnostic accuracy of the EUS-FNA group were 96.4% (27/28), 83.3% (10/12), 93.1% (27/29), 90.9% (10/11) and 92.5% (37/40), while those in the SpyGlass group were 100% (54/54), 75% (3/4), 98.2% (54/55), 100% (3/3) and 98.3% (57/58), respectively. The diagnostic accuracy rate in the SpyGlass group was higher than that in the EUS-FNA group; however, no significant difference was found between the two groups (P = 0.368). The diagnostic accuracy of evaluating specific cyst types in the EUS-FNA group was 85% (34/40), similar to that in the SpyGlass group (85.0% vs 84.5%, P = 0.944). CONCLUSION: SpyGlass seems less valuable for the diagnosis of PCLs when EUS and EUS-FNA have been performed by experienced endoscopists.

Clinical outcomes of endoscopic resection for the treatment of gastric gastrointestinal stromal tumors originating from the muscularis propria: a 7-year experience from a large tertiary center in China.

BACKGROUND AND AIMS: Few studies have evaluated the value of endoscopic resection (ER) for the treatment of gastric gastrointestinal (GI) stromal tumors (GISTs) originating from the muscularis propria (MP) in a large population, and no studies have evaluated risk factors for piecemeal resection. This study aimed to evaluate the efficacy and safety of ER for gastric GISTs in a real-world setting and to explore factors associated with piecemeal resection. METHODS: From January 2013 to December 2019, 185 patients with gastric GISTs originating from the MP were assessed. Clinicopathological and endoscopic data were collected and analyzed. Baseline characteristics of the en bloc resection and piecemeal resection groups were compared to evaluate predictive factors for piecemeal resection. RESULTS: There were 71 males and 114 females with a mean age of 57.0 ± 9.7 years. The mean size of GISTs was 15 mm (range 4-65 mm). A total of 123 were very low risk, 52 were low risk, and ten were moderate risk. In this study, 103 GISTs were treated with endoscopic submucosal excavation (ESE), 68 GISTs were treated with endoscopic full-thickness resection (EFR), and 14 GISTs were treated with submucosal tunneling ER (STER). Either en bloc resection or complete resection was achieved in 160 (86.5%) patients. No recurrence was noted during follow-up. Only five patients experienced minor complications, with a complication rate of 2.7%. Multivariate analysis demonstrated that size (odds ratio [OR] 1.060, 95% confidence interval [CI] 1.004-1.118; P = 0.035) and shape (OR 5.434, 95% CI 1.638-18.027; P = 0.006) were independent predictors of piecemeal resection. CONCLUSION: ER was effective and safe for the treatment of gastric GISTs originating from the MP. Piecemeal resection did not seem to affect the efficacy of ER, and no recurrence was noted during follow-up. Large size and irregular shape are risk factors related to piecemeal resection of ER.

Characterization of Copper/Zinc Superoxide Dismutase Activity on Phascolosoma esculenta (Sipuncula: Phascolosomatidea) and Its Protection from Oxidative Stress Induced by Cadmium.

Phascolosoma esculenta, an economically important species inhabiting the high tide areas of the intertidal zone, is particularly sensitive to water pollution. Considering its potential as a bioindicator, studies on the ecotoxicology of P. esculenta are imperative. The toxic effects of cadmium (Cd) were analyzed by exposing P. esculenta to different concentrations of Cd (6, 24, 96 mg/L). In this study, the changes in the antioxidative indexes of total superoxide dismutase (T-SOD), glutathione s-transferase (GST), reduced glutathione (GSH), and microscale malondialdehyde (MDA) were recorded. Copper/zinc superoxide dismutase (Cu/Zn SOD) is one of the most important free radical scavenging members. To reveal the antioxidative function of P. esculenta, an important member of the antioxidative system, designated Pe-Cu/Zn SOD, was cloned and analyzed. Phylogenic analysis revealed that Pe-Cu/Zn SOD was located in the invertebrate evolutionary branch of intracellular Cu/Zn SOD (icCu/Zn SOD). The quantitative real-time polymerase chain reaction results showed that Pe-Cu/Zn SOD messenger ribonucleic acid was widely expressed in all tissues examined. The highest expression levels in coelomic fluid after Cd exposure indicated its function in the stress response. Using a prokaryotic expression system, we obtained a Pe-Cu/Zn SOD recombinant protein, which enhanced the heavy metal tolerance of Escherichia coli. In vivo assays also confirmed that the Pe-Cu/Zn SOD recombinant protein had an antioxidative and free radical scavenging ability. A Cd toxicity experiment, in which purified Pe-Cu/Zn SOD protein was injected into the body cavities of P. esculenta, showed that the reactive oxygen species content in the coelomic fluid of the experimental group was significantly lower compared with the control group. These results suggest that Pe-Cu/Zn SOD played a role in Cd detoxification by chelating heavy metal ions and scavenging reactive oxygen free radicals, and that P. esculenta could be used as a bioindicator to evaluate heavy metal pollution.

Characterization of Mitochondrial Prohibitin in Opsariichthys bidens and Its Potential Functions in Spermatogenesis.

Spermatogenesis is the intricate and coordinated process by which spermatogonia develop into haploid differentiated spermatozoa. Mitochondria are essential for spermatogenesis, and prohibitin (PHB) is closely associated with mitochondrial structure and function during spermatogenesis. Although PHB has been implicated in spermatogenesis in some taxa, its roles in Opsariichthys bidens have not been determined. In this study, the expression patterns and potential functions of PHB in spermatogenesis in O. bidens were characterized using histological microscopic observations, PCR cloning, real-time quantitative PCR (qPCR), Western blotting (WB) and immunofluorescence (IF). The full-length cDNA of Ob-phb was 1500 bp encoding 271 amino acids. A sequence alignment demonstrated that the PHB protein is conserved among different animals. qPCR revealed that phb mRNA is widely distributed in O. bidens and highly expressed in the testes at stages IV and V. WB revealed that Ob-PHB is located in the mitochondria of testes. IF revealed the colocalization of PHB signals and mitochondria. Signals were detected around nuclei in spermatogonia and spermatocytes, gradually moving to the tail region during spermiogenesis, and finally aggregating in the midpiece. These results indicate that Ob-PHB was expressed in the mitochondria during spermatogenesis. In addition, this study proposed Ob-PHB may participate in the degradation of mitochondria and cell differentiation during spermatogenesis.