Genomic and transcriptomic studies on flavonoid biosynthesis in Lagerstroemia indica.

BACKGROUND: Lagerstroemia indica is a widely cultivated ornamental woody shrub/tree of the family Lythraceae that is used as a traditional medicinal plant in East Asia and Egypt. However, unlike other ornamental woody plants, its genome is not well-investigated, which hindered the discovery of the key genes that regulate important traits and the synthesis of bioactive compounds. RESULTS: In this study, the genomic sequences of L. indica were determined using several next-generation sequencing technologies. Altogether, 324.01 Mb sequences were assembled and 98.21% (318.21 Mb) of them were placed in 24 pseudo-chromosomes. The heterozygosity, repeated sequences, and GC residues occupied 1.65%, 29.17%, and 38.64% of the genome, respectively. In addition, 28,811 protein-coding gene models, 327 miRNAs, 552 tRNAs, 214 rRNAs, and 607 snRNAs were identified. The intra- and interspecies synteny and Ks analysis revealed that L. indica exhibits a hexaploidy. The co-expression profiles of the genes involved in the phenylpropanoid (PA) and flavonoid/anthocyanin (ABGs) pathways with the R2R3 MYB genes (137 members) showed that ten R2R3 MYB genes positively regulate flavonoid/anthocyanin biosynthesis. The colors of flowers with white, purple (PB), and deep purplish pink (DPB) petals were found to be determined by the levels of delphinidin-based (Dp) derivatives. However, the substrate specificities of LiDFR and LiOMT probably resulted in the different compositions of flavonoid/anthocyanin. In L. indica, two LiTTG1s (LiTTG1-1 and LiTTG1-2) were found to be the homologs of AtTTG1 (WD40). LiTTG1-1 was found to repress anthocyanin biosynthesis using the tobacco transient transfection assay. CONCLUSIONS: This study showed that the ancestor L. indica experienced genome triplication approximately 38.5 million years ago and that LiTTG1-1 represses anthocyanin biosynthesis. Furthermore, several genes such as LiDFR, LiOMTs, and R2R3 LiMYBs are related to anthocyanin biosynthesis. Further studies are required to clarify the mechanisms and alleles responsible for flower color development.

Evolution origin analysis and health risk assessment of groundwater environment in a typical mining area: Insights from water-rock interaction and anthropogenic activities.

Coal mining changes groundwater environment, results in deterioration of water quality and endangering human health in the mining area. However, the comprehensive study of groundwater evolution and its potential impact in mining area is still insufficient. In this study, 95 groundwater samples were collected from 2019 to 2020 in a typical mining area of China. Ion ratio coefficients, isotopic tracing technology, Entropy-weighted water quality index (EWQI) and human health risk assessment model (HHRA) were applicated to investigate the hydrochemical variation reasons, groundwater quality and its potential health risk in the study area. Results showed that the groundwater hydrochemical types changed from HCO3∙SO4-Ca∙Mg type to SO4-Ca∙Mg and SO4∙Cl-Ca∙Mg type. Water-rock interaction, agricultural activities, manure and sewage input, precipitation and evaporation controlled the groundwater hydrochemical composition. Groundwater quality showed a trend of fluctuation with an average EWQI of 59.23, 68.92, 63.75, 58.02 and 64.92, respectively. 91.6% of the water samples was fair and acceptable for drinking. The groundwater health risk of nitrate in the study area ranged from 0.03 to 17.80. Infants had the highest health risk and nitrate concentration was the most sensitive parameter. The results will present a comprehensive research of groundwater evolution and potential impacts through a typical mining area example. Thereby offering valuable insights into the influencing factors identification, hydrochemical processes evolution, protection and utilization of groundwater in global mining areas.

Ancient forest plants possess cytotoxic properties causing liver cancer HepG2 cell apoptosis.

Here, we collected 154 plant species in China ancient forests looking for novel efficient bioactive compounds for cancer treatments. We found 600 bioactive phyto-chemicals that induce apoptosis of liver cancer cell in vitro. First, we screen the plant extract's in vitro cytotoxicity inhibition of cancer cell growth using in vitro HepG2 cell lines and MTT cytotoxicity. The results from these initial MTT in vitro cytotoxicity tests show that the most efficient plants towards hepatoma cytoxicity is Cephalotaxus sinensis, mint bush (Elsholtzia stauntonii) and winged spindle tree (Euonymus alatus). We then used in cell-counting kit-8 (CCK-8) to further understand in vivo tumor growth using nude mice and GC-MS and LC-QTOF-MS to analyze the composition of compounds in the extracts. Extracted chemically active molecules analyzed by network pharmacology showed inhibition on the growth of liver cancer cells by acting on multiple gene targets, which is different from the currently used traditional drugs acting on only one target of liver cancer cells. Extracts from Cephalotaxus sinensis, mint bush (Elsholtzia stauntonii) and winged spindle tree (Euonymus alatus) induce apoptosis in hepatoma cancer cell line HepG2 with a killing rate of more than 83% and a tumor size decrease by 62-67% and a killing rate of only 6% of normal hepatocyte LO2. This study highlight efficient candidate species for cancer treatment providing a basis for future development of novel plant-based drugs to help meeting several of the UN SDGs and planetary health.

ERK/CREB and p38 MAPK/MMP14 Signaling Pathway Influences Spermatogenesis through Regulating the Expression of Junctional Proteins in Eriocheir sinensis Testis.

The hemolymph-testis barrier (HTB) is a reproduction barrier in Crustacea, guaranteeing the safe and smooth process of spermatogenesis, which is similar to the blood-testis barrier (BTB) in mammals. The MAPK signaling pathway plays an essential role in spermatogenesis and maintenance of the BTB. However, only a few studies have focused on the influence of MAPK on crustacean reproduction. In the present study, we knocked down and inhibited MAPK in Eriocheir sinensis. Increased defects in spermatogenesis were observed, concurrently with a damaged HTB. Further research revealed that es-MMP14 functions downstream of ERK and p38 MAPK and degrades junctional proteins (Pinin and ZO-1); es-CREB functions in the ERK cascade as a transcription factor of ZO-1. In addition, when es-MMP14 and es-CREB were deleted, the defects in HTB and spermatogenesis aligned with abnormalities in the MAPK. However, JNK impacts the integrity of the HTB by changing the distribution of intercellular junctions. In summary, the MAPK signaling pathway maintains HTB integrity and spermatogenesis through es-MMP14 and es-CREB, which provides insights into the evolution of gene function during barrier evolution.

Central venous oxygen saturation changes as a reliable predictor of the change of CI in septic shock: To explore potential influencing factors.

PURPOSE: Assessing fluid responsiveness relying on central venous oxygen saturation (ScvO2) yields varied outcomes across several studies. This study aimed to determine the ability of the change in ScvO2 (ΔScvO2) to detect fluid responsiveness in ventilated septic shock patients and potential influencing factors. METHODS: In this prospective, single-center study, all patients conducted from February 2023 to January 2024 received fluid challenge. Oxygen consumption was measured by indirect calorimetry, and fluid responsiveness was defined as an increase of cardiac index (CI) ≥ 10% measured by transthoracic echocardiography. Multivariate linear regression analysis was conducted to evaluate the impact of oxygen consumption, arterial oxygen saturation, CI, and hemoglobin on ScvO2 and its change before and after fluid challenge. RESULTS: Among 49 patients (31 men, aged (59 ± 18) years), 27 were responders. The patients had an acute physiology and chronic health evaluation II score of 24 ± 8, a sequential organ failure assessment score of 11 ± 4, and a blood lactate level of (3.2 ± 3.1) mmol/L at enrollment. After the fluid challenge, the ΔScvO2 (mmHg) in the responders was greater than that in the non-responders (4 ± 6 vs. 1 ± 3, p = 0.019). Multivariate linear regression analysis suggested that CI was the only independent influencing factor of ScvO2, with R2 = 0.063, p = 0.008. After the fluid challenge, the change in CI became the only contributing factor to ΔScvO2 (R2 = 0.245, p < 0.001). ΔScvO2 had a good discriminatory ability for the responders and non-responders with a threshold of 4.4% (area under the curve = 0.732, p = 0.006). CONCLUSION: ΔScvO2 served as a reliable surrogate marker for ΔCI and could be utilized to assess fluid responsiveness, given that the change of CI was the sole contributing factor to the ΔScvO2. In stable hemoglobin conditions, the absolute value of ScvO2 could serve as a monitoring indicator for adequate oxygen delivery independent of oxygen consumption.

mTORC1/rpS6 and mTORC2/PKC regulate spermatogenesis through Arp3-mediated actin microfilament organization in Eriocheir sinensis.

Mammalian target of rapamycin (mTOR) is a crucial signaling protein regulating a range of cellular events. Numerous studies have reported that the mTOR pathway is related to spermatogenesis in mammals. However, its functions and underlying mechanisms in crustaceans remain largely unknown. mTOR exists as two multimeric functional complexes termed mTOR complex 1 (mTORC1) and mTORC2. Herein, we first cloned ribosomal protein S6 (rpS6, a downstream molecule of mTORC1) and protein kinase C (PKC, a downstream effector of mTORC2) from the testis of Eriocheir sinensis. The dynamic localization of rpS6 and PKC suggested that both proteins may be essential for spermatogenesis. rpS6/PKC knockdown and Torin1 treatment led to defects in spermatogenesis, including germ cell loss, retention of mature sperm and empty lumen formation. In addition, the integrity of the testis barrier (similar to the blood-testis barrier in mammals) was disrupted in the rpS6/PKC knockdown and Torin1 treatment groups, accompanied by changing in expression and distribution of junction proteins. Further study demonstrated that these findings may result from the disorganization of filamentous actin (F-actin) networks, which were mediated by the expression of actin-related protein 3 (Arp3) rather than epidermal growth factor receptor pathway substrate 8 (Eps8). In summary, our study illustrated that mTORC1/rpS6 and mTORC2/PKC regulated spermatogenesis via Arp3-mediated actin microfilament organization in E. sinensis.

Complete genome analysis of pathogenic Metschnikowia bicuspidata strain MQ2101 isolated from diseased ridgetail white prawn, Exopalaemon carinicauda.

BACKGROUND: Metschnikowia bicuspidata is a pathogenic yesst that can cause disease in many different economic aquatic animal species. In recent years, there was a new disease outbreak in ridgetail white prawn (Exopalaemon carinicauda) in coastal areas of Jiangsu Province China that was referred to as zombie disease by local farmers. The pathogen was first isolated and identified as M. bicuspidata. Although the pathogenicity and pathogenesis of this pathogen in other animals have been reported in some previous studies, research on its molecular mechanisms is still very limited. Therefore, a genome-wide study is necessary to better understand the physiological and pathogenic mechanisms of M. bicuspidata. RESULT: In this study, we obtained a pathogenic strain, MQ2101, of M. bicuspidata from diseased E. carinicauda and sequenced its whole genome. The size of the whole genome was 15.98 Mb, and it was assembled into 5 scaffolds. The genome contained 3934 coding genes, among which 3899 genes with biological functions were annotated in multiple underlying databases. In KOG database, 2627 genes were annotated, which were categorized into 25 classes including general function prediction only, posttranslational modification, protein turnover, chaperones, and signal transduction mechanisms. In KEGG database, 2493 genes were annotated, which were categorized into five classes, including cellular processes, environmental information processing, genetic information processing, metabolism and organismal systems. In GO database, 2893 genes were annotated, which were mainly classified in cell, cell part, cellular processes and metabolic processes. There were 1055 genes annotated in the PHI database, accounting for 26.81% of the total genome, among which 5 genes were directly related to pathogenicity (identity ≥ 50%), including hsp90, PacC, and PHO84. There were also some genes related to the activity of the yeast itself that could be targeted by antiyeast drugs. Analysis based on the DFVF database showed that strain MQ2101 contained 235 potential virulence genes. BLAST searches in the CAZy database showed that strain MQ2101 may have a more complex carbohydrate metabolism system than other yeasts of the same family. In addition, two gene clusters and 168 putative secretory proteins were predicted in strain MQ2101, and functional analysis showed that some of the secretory proteins may be directly involved in the pathogenesis of the strain. Gene family analysis with five other yeasts revealed that strain MQ2101 has 245 unique gene families, including 274 genes involved in pathogenicity that could serve as potential targets. CONCLUSION: Genome-wide analysis elucidated the pathogenicity-associated genes of M. bicuspidate while also revealing a complex metabolic mechanism and providing putative targets of action for the development of antiyeast drugs for this pathogen. The obtained whole-genome sequencing data provide an important theoretical basis for transcriptomic, proteomic and metabolic studies of M. bicuspidata and lay a foundation for defining its specific mechanism of host infestation.

Comparing biweekly single-dose actinomycin D with multiday methotrexate therapy for low-risk gestational trophoblastic neoplasia (FIGO Score 0-4): study protocol for a prospective, multicentre, randomized trial.

BACKGROUND: Single-agent chemotherapy using methotrexate or actinomycin D is the first-line treatment for patients with low-risk gestational trophoblastic neoplasia. Various methotrexate-based and actinomycin D-based single-agent regimens can be used. However, there is insufficient evidence to determine the superior regimen. To guide doctors in selecting a single-agent chemotherapy regimen for patients with low-risk gestational trophoblastic neoplasia, we will compare two regimens. METHODS: We will conduct a multicentre, randomized, prospective clinical trial. Selected low-risk gestational trophoblastic neoplasia patients (FIGO score 0-4) will be randomized 1:1 to a biweekly single-dose actinomycin D group or a multiday methotrexate therapy group. The actinomycin D group will receive IV pulse actinomycin D (1.25 mg/m2) every 14 days, and the methotrexate group will receive methotrexate (50 mg) intramuscularly on days 1, 3, 5, and 7 (4 doses per cycle) and leucovorin (15 mg) intramuscularly on days 2, 4, 6, and 8. This process will be repeated every 14 days. The primary endpoints will include the complete remission rate by single-agent therapy and the overall complete remission rate. The secondary endpoints will include the duration needed to achieve complete remission after single-agent chemotherapy, number of courses needed to achieve complete remission after single-agent chemotherapy, incidence and severity of adverse effects, effects on menstrual conditions and ovarian function based on the anti-Mullerian hormone level, and patient-reported quality of life. DISCUSSION: Previous clinical trials comparing biweekly single-dose actinomycin D with multiday methotrexate therapy for treating low-risk gestational trophoblastic neoplasia patients failed to meet the expected case number. Through this multicentre study, the complete remission ratio and efficacy difference between biweekly single-dose actinomycin D and multiday methotrexate therapy will be obtained. This study will also provide the basis for formulating a preferred regimen for treating patients with low-risk gestational trophoblastic neoplasia. TRIAL REGISTRATION: ClinicalTrials.gov: NCT04562558, Registered on 13 September 2020 (Protocol version 2020-9-24, version 1.0).

Tissue oxygen saturation is predictive of lactate clearance in patients with circulatory shock.

BACKGROUND: Tissue oxygen saturation (StO2) decrease could appear earlier than lactate alteration. However, the correlation between StO2 and lactate clearance was unknown. METHODS: This was a prospective observational study. All consecutive patients with circulatory shock and lactate over 3 mmol/L were included. Based on the rule of nines, a BSA (body surface area) weighted StO2 was calculated from four sites of StO2 (masseter, deltoid, thenar and knee). The formulation was as follows: masseter StO2 × 9% + (deltoid StO2 + thenar StO2) × (18% + 27%)/ 2 + knee StO2 × 46%. Vital signs, blood lactate, arterial and central venous blood gas were measured simultaneously within 48 h of ICU admission. The predictive value of BSA-weighted StO2 on 6-hour lactate clearance > 10% since StO2 initially monitored was assessed. RESULTS: A total of 34 patients were included, of whom 19 (55.9%) had a lactate clearance higher than 10%. The mean SOFA score was lower in cLac ≥ 10% group compared with cLac < 10% group (11 ± 3 vs. 15 ± 4, p = 0.007). Other baseline characteristics were comparable between groups. Compared to non-clearance group, StO2 in deltoid, thenar and knee were significantly higher in clearance group. The area under the receiver operating curves (AUROC) of BSA-weighted StO2 for prediction of lactate clearance (0.92, 95% CI [Confidence Interval] 0.82-1.00) was significantly higher than StO2 of masseter (0.65, 95% CI 0.45-0.84; p < 0.01), deltoid (0.77, 95% CI 0.60-0.94; p = 0.04), thenar (0.72, 95% CI 0.55-0.90; p = 0.01), and similar to knee (0.87, 0.73-1.00; p = 0.40), mean StO2 (0.85, 0.73-0.98; p = 0.09). Additionally, BSA-weighted StO2 model had continuous net reclassification improvement (NRI) over the knee StO2 and mean StO2 model (continuous NRI 48.1% and 90.2%, respectively). The AUROC of BSA-weighted StO2 was 0.91(95% CI 0.75-1.0) adjusted by mean arterial pressure and norepinephrine dose. CONCLUSIONS: Our results suggested that BSA-weighted StO2 was a strong predictor of 6-hour lactate clearance in patients with shock.

Pituitary adenoma with oculomotor cistern extension: membranous anatomy and clinical application.

BACKGROUND: The anatomical basis of pituitary adenomas (PAs) with oculomotor cistern (OC) extension as a growth corridor is overlooked in the literature. In this paper, the authors use the technique of epoxy sheet plastination to study the membranous structure of the OC and validate the results by retrospective analysis of patients with OC extension. METHODS: Eighteen specimens were used to study the membranous anatomy surrounding the OC using the epoxy sheet plastination technique. Thirty-four patients with OC extension were retrospectively reviewed. RESULTS: The OC consisted of two thin membranous layers. The inner layer was extended by the arachnoid layer from the posterior fossa, and the lateral layer consisted of the dura mater sinking from the roof of the cavernous sinus. The oculomotor nerve is more likely to displace with a superolateral trajectory due to the weakness of the posterior dura and the relatively large space in the medial and posterior trajectories, which is consistent with the intraoperative observations. Among the anatomical factors that affect the PA by OC extension, we found that the relative position of the internal carotid artery (ICA) and posterior clinoid process may lead to the narrowing of the OC. Of 34 cases, 28 patients achieved total resection. Among 24 preoperative patients with oculomotor nerve palsy, 16 cases were relieved to varying degrees postoperatively. There was no ICA injury or severe intracranial infection found in any of the patients. CONCLUSIONS: Extension into the OC is influenced by two anatomical factors: a weak point in the dura in the posterior OC and a potential space beyond this region of the dura. Meticulous knowledge of the membranous anatomy in endoscopic endonasal surgery is required to safely and effectively resect PA with OC extension.

mTORC1/C2 regulate spermatogenesis in Eriocheir sinensis via alterations in the actin filament network and cell junctions.

Spermatogenesis is a finely regulated process of germ cell proliferation and differentiation that leads to the production of sperm in seminiferous tubules. Although the mammalian target of rapamycin (mTOR) signaling pathway is crucial for spermatogenesis in mammals, its functions and molecular mechanisms in spermatogenesis remain largely unknown in nonmammalian species, particularly in Crustacea. In this study, we first identified es-Raptor (the core component of mTOR complex 1) and es-Rictor (the core component of mTOR complex 2) from the testis of Eriocheir sinensis. Dynamic localization of es-Raptor and es-Rictor implied that these proteins were indispensable for the spermatogenesis of E. sinensis. Furthermore, es-Raptor and es-Rictor knockdown results showed that the mature sperm failed to be released, causing almost empty lumens in the testis. We investigated the reasons for these effects and found that the actin-based cytoskeleton was disrupted in the knockdown groups. In addition, the integrity of the testis barrier (similar to the blood-testis barrier in mammals) was impaired and affected the expression of cell junction proteins. Further study revealed that es-Raptor and es-Rictor may regulate spermatogenesis via both mTORC1- and mTORC2-dependent mechanisms that involve es-rpS6 and es-Akt/es-PKC, respectively. Moreover, to explore the testis barrier in E. sinensis, we established a cadmium chloride (CdCl2)-induced testis barrier damage model as a positive control. Morphological and immunofluorescence results were similar to those of the es-Raptor and es-Rictor knockdown groups. Altogether, es-Raptor and es-Rictor were important for spermatogenesis through maintenance of the actin filament network and cell junctions in E. sinensis.

Follicle-stimulating hormone signaling in Sertoli cells: a licence to the early stages of spermatogenesis.

Follicle-stimulating hormone signaling is essential for the initiation and early stages of spermatogenesis. Follicle-stimulating hormone receptor is exclusively expressed in Sertoli cells. As the only type of somatic cell in the seminiferous tubule, Sertoli cells regulate spermatogenesis not only by controlling their own number and function but also through paracrine actions to nourish germ cells surrounded by Sertoli cells. After follicle-stimulating hormone binds to its receptor and activates the follicle-stimulating hormone signaling pathway, follicle-stimulating hormone signaling will establish a normal Sertoli cell number and promote their differentiation. Spermatogonia pool maintenance, spermatogonia differentiation and their entry into meiosis are also positively regulated by follicle-stimulating hormone signaling. In addition, follicle-stimulating hormone signaling regulates germ cell survival and limits their apoptosis. Our review summarizes the aforementioned functions of follicle-stimulating hormone signaling in Sertoli cells. We also describe the clinical potential of follicle-stimulating hormone treatment in male patients with infertility. Furthermore, our review may be helpful for developing better therapies for treating patients with dysfunctional follicle-stimulating hormone signaling in Sertoli cells.

Lessons on Drug Development: A Literature Review of Challenges Faced in Nonalcoholic Fatty Liver Disease (NAFLD) Clinical Trials.

NAFLD is the most common chronic liver disease worldwide, occurring in both obese and lean patients. It can lead to life-threatening liver diseases and nonhepatic complications, such as cirrhosis and cardiovascular diseases, that burden public health and the health care system. Current care is weight loss through diet and exercise, which is a challenging goal to achieve. However, there are no FDA-approved pharmacotherapies for NAFLD. This review thoroughly examines the clinical trial findings from 22 drugs (Phase 2 and above) and evaluates the future direction that trials should take for further drug development. These trialed drugs can broadly be categorized into five groups-hypoglycemic, lipid-lowering, bile-pathway, anti-inflammatory, and others, which include nutraceuticals. The multitude of challenges faced in these yet-to-be-approved NAFLD drug trials provided insight into a few areas of improvement worth considering. These include drug repurposing, combinations, noninvasive outcomes, standardization, adverse event alleviation, and the need for precision medicine with more extensive consideration of NAFLD heterogenicity in drug trials. Understandably, every evolution of the drug development landscape lies with its own set of challenges. However, this paper believes in the importance of always learning from lessons of the past, with each potential improvement pushing clinical trials an additional step forward toward discovering appropriate drugs for effective NAFLD management.

Kinesin 12 (KIF15) contributes to the development and tumorigenicity of prostate cancer.

In Asia, prostate cancer is becoming a growing concern, impacting both socially and economically, compared with what is seen in western countries. Hence, it is essential to know the mechanisms associated with the development and tumorigenesis of PCa for primary diagnosis, risk management, and development of therapy strategies against PCa. Kinesin family member 15 (KIF15), a kinesin family member, is a plus-end-directed kinesin that functions to form bipolar spindles. There is emerging evidence indicating that KIF15 plays a significant role in several malignancies, such as pancreatic cancer, hepatocellular carcinoma, lung adenocarcinoma, and breast cancer. Still, the function of KIF15 remains unclear in prostate cancer. Here, we study the functional importance of KIF15 in the tumorigenesis of PCa. The bioinformatic analysis from PCa patients revealed high KIF15 expression compared to normal prostate tissues. High expression hinting at a possible functional role of KIF15 in regulating cell proliferation of PCa, which was demonstrated by both in vitro and in vivo assays. Downregulation of KIF15 silenced the expression of CDK2, p-RB, and Cyclin D1 and likewise blocked the cells at the G1 stage of the cell cycle. In addition, KIF15 downregulation inhibited MEK-ERK signaling by significantly silencing p-ERK and p-MEK levels. In conclusion, this study confirmed the functional significance of KIF15 in the growth and development of prostate cancer and could be a novel therapeutic target for the treatment of PCa.

PIWIs maintain testis apoptosis to remove abnormal germ cells in Eriocheir sinensis.

PIWI proteins play important roles in germline development in the mammals. However, the functions of PIWIs in crustaceans remain unknown. In the present study, we identified three Piwis from the testis of Eriocheir sinensis (E. sinensis). Three Piwi genes encoded proteins with typical features of PIWI subfamilies and were highly expressed in the testis. Three PIWIs could be detected in the cytoplasm of spermatocytes and spermatids, while in spermatozoa, we could only detect PIWI1 and PIWI3 in the nucleus. The knockdown of PIWIs by dsRNA significantly affected the formation of the nuclei in spermatozoa, which resulted in deviant and irregular nuclei. PIWI defects significantly inhibited the apoptosis of abnormal germ cells through the caspase-dependent apoptosis pathway and p53 pathway. Knockdown of PIWIs inhibited the expression of caspase (Casp) 3, 7, 8, and p53 without affecting Bcl2 (B-cell lymphoma gene 2), Bax (B-cell lymphoma-2-associated X), and BaxI (B-cell lymphoma-2-associated X inhibitor), which further significantly increased abnormal spermatozoa in the knockdown-group crabs. These results show a new role of PIWI proteins in crustaceans that is different from that in mammals. In summary, PIWIs play roles in the formation of the germline nucleus and can maintain apoptosis in abnormal germ cells to remove abnormal germ cells in E. sinensis.

KIF3A regulates the Wnt/ß-catenin pathway via transporting ß-catenin during spermatogenesis in Eriocheir sinensis.

The Wnt/ß-catenin pathway participates in many important physiological events such as cell proliferation and differentiation in the male reproductive system. We found that Kinesin-2 motor KIF3A is highly expressed during spermatogenesis in Eriocheir sinensis; it may potentially promote the intracellular transport of cargoes in this process. However, only a few studies have focused on the relationship between KIF3A and the Wnt/ß-catenin pathway in the male reproductive system of decapod crustaceans. In this study, we cloned and characterized the CDS of ß-catenin in E. sinensis for the first time. Fluorescence in situ hybridization and immunofluorescence results showed the colocalization of Es-KIF3A and Es-ß-catenin at the mRNA and the protein level respectively. To further explore the regulatory function of Es-KIF3A to the Wnt/ß-catenin pathway, the es-kif3a was knocked down by double-stranded RNA (dsRNA) in vivo and in primary cultured cells in testes of E. sinensis. Results showed that the expression of es-ß-catenin and es-dvl were decreased in the es-kif3a knockdown group. The protein expression level of Es-ß-catenin was also reduced and the location of Es-ß-catenin was changed from nucleus to cytoplasm in the late stage of spermatogenesis when es-kif3a was knocked down. Besides, the co-IP result demonstrated that Es-KIF3A could bind with Es-ß-catenin. In summary, this study indicates that Es-KIF3A can positively regulate the Wnt/ß-catenin pathway during spermatogenesis and Es-KIF3A can bind with Es-ß-catenin to facilitate the nuclear translocation of Es-ß-catenin.

Reproductive outcomes after floxuridine-based regimens for gestational trophoblastic neoplasia: A retrospective cohort study in a national referral center in China.

OBJECTIVE: To analyze the reproductive outcomes of gestational trophoblastic neoplasia (GTN) patients who were cured only by floxuridine-based regimens. METHODS: This was a retrospective analysis of 464 patients who were treated with only floxuridine-based regimens at Peking Union Medical College Hospital between January 2002 and December 2013 and retained their reproductive ability. Their reproductive outcomes were analyzed. The factors affecting pregnancy intention were identified by logistic regression. RESULTS: Of the 464 patients (average age, 28.0 ± 5.7 years; median follow-up = 85 months), the livebirth rate was 72.2%, while the rates of spontaneous abortion, induced abortion and ectopic pregnancy were 9.2% (n = 41), 8.7% (n = 39) and 1.8% (n = 8), respectively. The GTN recurrence rate was 2.1%. The time from chemotherapy completion to first conception in the induced abortion group was significantly shorter than those in spontaneous abortion, full-term/premature, and ectopic pregnancy groups (P ≤ 0.001, <0.001, =0.015, respectively). The logistic analysis showed that the age at onset of GTN (OR = 0.899, 95% CI 0.851-0.951, P < 0.001), parity at onset of GTN (parity = 1, OR = 0.123, 95% CI 0.068-0.225, P < 0.001; parity = 2-3, OR = 0.058, 95% CI 0.014-0.232, P < 0.001) and interval from the index pregnancy to chemotherapy were independent factors affecting pregnancy intention. Among the 36 pregnancies occurring within 12 months postchemotherapy, only one choriocarcinoma occurred, and 20 culminated in induced abortions (55.6%). CONCLUSIONS: After floxuridine-based chemotherapy, the pregnancy rate of GTN patients after fertility-preserving treatment is comparable to that of the normal population. Pregnancy losses within one year after chemotherapy completion are mainly caused by induced abortion.

SOX-mediated molecular crosstalk during the progression of tumorigenesis.

SOX family transcription factor has emerged as a double-edged sword relating to tumorigenesis and metastasis. Multiple studies have revealed different expression patterns and contradictory roles of SOX factors in the tumor initiation and progression. The aberrant expression of SOX factors is regulated by copy number alteration, methylation modulation, microRNAs, transcription factors and post-translational modification. This review summarizes the role of SOX factors in molecular interactions and signaling pathways during different steps of carcinogenesis, such as CSCs stemness maintenance, EMT occurrence, cell invasion, cell proliferation and apoptosis. The Wnt signaling pathway is also shown to provide vital intermediate signaling transduction. We believe that SOX family proteins may be used as prognostic markers for human clinical therapy, and novel therapy strategies targeting SOX factors should be explored in future clinical applications.

Sry and SoxE genes: How they participate in mammalian sex determination and gonadal development?

In mammals, sex determination defines the differentiation of the bipotential genital ridge into either testes or ovaries. Sry, the mammalian Y-chromosomal testis-determining gene, is a master regulator of male sex determination. It acts to switch the undifferentiated genital ridge towards testis development, triggering the adoption of a male fate. Sry initiates a cascade of gene networks through the direct regulation of Sox9 expression and promotes supporting cell differentiation, Leydig cell specification, vasculature formation and testis cord development. In the absence of Sry, alternative genetic cascades, including female sex-determining genes RSPO1, Wnt4/ß-catenin and Foxl2, are involved in the formation of female genitalia and the maintenance of female ovarian development. The mutual antagonisms between male and female sex-determining pathways are crucial in not just the initiation but also the maintenance of the somatic sex of the gonad throughout the organism's lifetime. Any imbalances in above sex-determining genes can cause disorders of sex development in humans and mice. In this review, we provide a detailed summary of the expression profiles, biochemical properties and developmental functions of Sry and SoxE genes in embryonic testis development and adult gonadal development. We also briefly summarize the dedicate balances between male and female sex-determining genes in mammalian sex development, with particular highlights on the molecular actions of Sry and Sox9 transcription factors.

Molecular mechanisms of kinesin-14 motors in spindle assembly and chromosome segregation.

During eukaryote cell division, molecular motors are crucial regulators of microtubule organization, spindle assembly, chromosome segregation and intracellular transport. The kinesin-14 motors are evolutionarily conserved minus-end-directed kinesin motors that occur in diverse organisms from simple yeasts to higher eukaryotes. Members of the kinesin-14 motor family can bind to, crosslink or slide microtubules and, thus, regulate microtubule organization and spindle assembly. In this Commentary, we present the common subthemes that have emerged from studies of the molecular kinetics and mechanics of kinesin-14 motors, particularly with regard to their non-processive movement, their ability to crosslink microtubules and interact with the minus- and plus-ends of microtubules, and with microtubule-organizing center proteins. In particular, counteracting forces between minus-end-directed kinesin-14 and plus-end-directed kinesin-5 motors have recently been implicated in the regulation of microtubule nucleation. We also discuss recent progress in our current understanding of the multiple and fundamental functions that kinesin-14 motors family members have in important aspects of cell division, including the spindle pole, spindle organization and chromosome segregation.