Complete chloroplast genome and comparison of herbicides toxicity on Aeschynomene indica (Leguminosae) in upland direct-seeding paddy field.

BACKGROUND: Indian jointvetch (Aeschynomene indica) is a common and pernicious weed found in the upland direct-seeding rice fields in the lower reaches of the Yangtze River in China. However, there are few reports on the degree of harm, genetic characteristics, and management methods of this weed. The purpose of this study is to clarify the harm of Indian jointvetch to upland direct-seeding rice, analyze the genetic characteristics of this weed based on chloroplast genomics and identify its related species, and screen herbicides that are effective in managing this weed in upland direct-seeding rice fields. RESULTS: In a field investigation in upland direct-seeding rice paddies in Shanghai and Jiangsu, we determined that the plant height and maximum lateral distance of Indian jointvetch reached approximately 134.2 cm and 57.9 cm, respectively. With Indian jointvetch present at a density of 4/m2 and 8/m2, the yield of rice decreased by approximately 50% and 70%, respectively. We further obtained the first assembly of the complete chloroplast (cp.) genome sequence of Indian jointvetch (163,613 bp). There were 161 simple sequence repeats, 166 long repeats, and 83 protein-encoding genes. The phylogenetic tree and inverted repeat region expansion and contraction analysis based on cp. genomes demonstrated that species with closer affinity to A. indica included Glycine soja, Glycine max, and Sesbania cannabina. Moreover, a total of 3281, 3840, and 3838 single nucleotide polymorphisms were detected in the coding sequence regions of the cp. genomes of S. cannabina voucher IBSC, G. soja, and G. max compared with the A. indica sequence, respectively. A greenhouse pot experiment indicated that two pre-emergence herbicides, saflufenacil and oxyfluorfen, and two post-emergence herbicides, florpyrauxifen-benzyl and penoxsulam, can more effectively manage Indian jointvetch than other common herbicides in paddy fields. The combination of these two types of herbicides is recommended for managing Indian jointvetch throughout the entire growth period of upland direct-seeding rice. CONCLUSIONS: This study provides molecular resources for future research focusing on the identification of the infrageneric taxa, phylogenetic resolution, and biodiversity of Leguminosae plants, along with recommendations for reliable management methods to control Indian jointvetch.

Microcystin-LR accelerates follicular atresia in mice via JNK-mediated adherent junction damage of ovarian granulosa cells.

Microcystin-LR (MC-LR), one of aquatic environmental contaminants with reproductive toxicity produced by cyanobacterial blooms, but its toxic effects and mechanisms on the ovary are not fully understood. Here, proteomic techniques and molecular biology experiments were performed to study the potential mechanism of MC-LR-caused ovarian toxicity. Results showed that protein expression profile of ovarian granulosa cells (KK-1) was changed by 17 µg/mL MC-LR exposure. Comparing with the control group, 118 upregulated proteins as well as 97 downregulated proteins were identified in MC-LR group. Function of differentially expressed proteins was found to be enriched in pathways related to adherent junction, such as cadherin binding, cell-cell junction, cell adhesion and focal adherens. Furthermore, in vitro experiments, MC-LR significantly downregulated the expression levels of proteins associated with adherent junction (ß-catenin, N-cadherin, and α-catenin) as well as caused cytoskeletal disruption in KK-1 cells (P < 0.05), indicating that the adherent junction was damaged. Results of in vivo experiments have shown that after 14 days of acute MC-LR exposure (40 µg/kg), damaged adherent junction and an increased number of atretic follicles were observed in mouse ovaries. Moreover, MC-LR activated JNK, an upstream regulator of adherent junction proteins, in KK-1 cells and mouse ovarian tissues. In contrast, JNK inhibition alleviated MC-LR-induced adherent junction damage in vivo and in vitro, as well as the number of atretic follicles. Taken together, findings from the present study indicated that JNK is involved in MC-LR-induced granulosa cell adherent junction damage, which accelerated follicular atresia. Our study clarified a novel mechanism of MC-LR-caused ovarian toxicity, providing a theoretical foundation for protecting female reproductive health from environmental pollutants.

PI3K/AKT/mTOR pathway mediated-cell cycle dysregulation contribute to malignant proliferation of mouse spermatogonia induced by microcystin-leucine arginine.

Environmental cyanotoxin exposure may be a trigger of testicular cancer. Activation of PI3K/AKT/mTOR signaling pathway is the critical molecular event in testicular carcinogenesis. As a widespread cyanotoxin, microcystin-leucine arginine (MC-LR) is known to induce cell malignant transformation and tumorigenesis. However, the effects of MC-LR on the regulatory mechanism of PI3K/AKT/mTOR pathway in seminoma, the most common testicular tumor, are unknown. In this study, mouse spermatogonia cell line (GC-1) and nude mice were used to investigate the effects and mechanisms of MC-LR on the malignant transformation of spermatogonia by nude mouse tumorigenesis assay, cell migration invasion assay, western blot, and cell cycle assay, and so forth. The results showed that, after continuous exposure to environmentally relevant concentrations of MC-LR (20 nM) for 35 generations, the proliferation, migration, and invasion abilities of GC-1 cells were increased by 120%, 340%, and 370%, respectively. In nude mice, MC-LR-treated GC-1 cells formed tumors with significantly greater volume (0.998 ± 0.768 cm3 ) and weight (0.637 ± 0.406 g) than the control group (0.067 ± 0.039 cm3 ; 0.094 ± 0.087 g) (P < .05). Furthermore, PI3K inhibitor Wortmannin inhibited the PI3K/AKT/mTOR pathway and its downstream proteins (c-MYC, CDK4, CCND1, and MMP14) activated by MC-LR. Blocking PI3K alleviated MC-LR-induced cell cycle disorder and malignant proliferation, migration and invasive of GC-1 cells. Altogether, our findings suggest that MC-LR can induce malignant transformation of mouse spermatogonia, and the PI3K/AKT/mTOR pathway-mediated cell cycle dysregulation may be an important target for malignant proliferation. This study provides clues to further reveal the etiology and pathogenesis of seminoma.

Long-term exposure to low concentrations of MC-LR induces blood-testis barrier damage through the RhoA/ROCK pathway.

Microcystin-leucine arginine (MC-LR), an emerging water pollutant, produced by cyanobacteria, has an acute testicular toxicity. However, little is known about the chronic toxic effects of MC-LR exposure on the testis at environmental concentrations and the underlying molecular mechanisms. In this study, C57BL/6 J mice were exposed to different low concentrations of MC-LR for 6, 9 and 12 months. The results showed that MC-LR could cause testis structure loss, cell abscission and blood-testis barrier (BTB) damage. Long-term exposure of MC-LR also activated RhoA/ROCK pathway, which was accompanied by the rearrangement of α-Tubulin. Furthermore, MC-LR reduced the levels of the adherens junction proteins (N-cadherin and ß-catenin) and the tight junction proteins (ZO-1 and Occludin) in a dose- and time-dependent way, causing BTB damage. MC-LR also reduced the expressions of Occludin, ZO-1, ß-catenin, and N-cadherin in TM4 cells, accompanied by a disruption of cytoskeletal proteins. More importantly, the RhoA inhibitor Rhosin ameliorated these MC-LR-induced changes. Together, these new findings suggest that long-term exposure to MC-LR induces BTB damage through RhoA/ROCK activation: involvement of tight junction and adherens junction changes and cytoskeleton disruption. This study highlights a new mechanism for MC-LR-induced BTB disruption and provides new insights into the cause and treatment of BTB disruption.

The optimization of ligature/bone defect-induced periodontitis model in rats.

The destruction of alveolar bone is a crucial manifestation of severe chronic periodontitis, which stem cell-based bioengineered therapies are expected to cure. Therefore, a cost-effective, reproducible, quantifiability and easier to administrate animal model that mimics human periodontitis is of great importance for further endeavor. In this study, we created periodontitis rat models in silk ligation group, bone defect group and bone defect/silk ligation group, respectively. Obvious periodontal inflammation but slight alveolar bone resorption was observed in the ligation group, while surgical trauma was not robust enough to continually worsen the constructed bone defect area in the bone defect group. In the bone defect/ligature group, significant and stable periodontal inflammation was the most enduring with similar evolving pathological patterns of human periodontitis. It also exhibited enhanced clinical similarity and confirmed its superiority in quantitativeness. The present rat model is the first study to reproduce a pathological process similar to human periodontitis with reliable stability and repeatability, manifesting a priority to previous methods. Day 9-12 is the best time for reproducing severe periodontitis syndromes with vertical bone resorption in this model.

Comparative Analysis of Whole Chloroplast Genomes of Three Common Species of Echinochloa (Gramineae) in Paddy Fields.

Echinochloa crus-galli var. crus-galli, E. crus-galli var. zelayensis, and E. glabrescens, morphologically similar at the seedling stage, are the most pernicious barnyard grass species in paddy fields worldwide. Chloroplast (cp) genomes could be conducive to their identification. In this study, we assembled the complete cp genome sequences of Echinochloa crus-galli var. crus-galli (139,856 bp), E. crus-galli var. zelayensis (139,874 bp), and E. glabrescens (139,874 bp), which exhibited a typical circular tetramerous structure, large and small single-copy regions, and a pair of inverted repeats. In Echinochloa crus-galli var. crus-galli, there were 136 simple sequence (SSRs) and 62 long (LRs) repeats, and in the other two species, 139 SSRs and 68 LRs. Each cp genome contains 92 protein-encoding genes. In Echinochloa crus-galli var. crus-galli and E. glabrescens, 321 and 1 single-nucleotide polymorphisms were detected compared to Echinochloa crus-galli var. zelayensis. IR expansion and contraction revealed small differences between the three species. The phylogenetic tree based on cp genomes demonstrated the phylogenetic relationship between ten barnyard grass species and other common Gramineae plants, showing new genetic relationships of the genus Echinochloa. This study provides valuable information on cp genomes, useful for identifying and classifying the genus Echinochloa and studying its phylogenetic relationships and evolution.

MicroRNA-122 overexpression promotes apoptosis and tumor suppressor gene expression induced by microcystin-leucine arginine in mouse liver.

Microcystin-leucine arginine (MC-LR), an important hepatoxin, has the effect of promoting hepatocarcinogenesis. MicroRNA-122 (miR-122), an important tumor suppressor in liver, plays an important role in promoting cell apoptosis. Previous studies found that the expression of miR-122 was reduced after MC-LR exposure in liver. In this study, C57BL/6 mice were exposed to saline, negative control agomir, and MC-LR with or without miR-122 agomir transfection. The results indicated that MC-LR promoted the expressions of tumor suppressor genes and decreased the expressions of anti-apoptotic proteins B cell lymphoma-2 (Bcl-2) and Bcl-2-like 2 (Bcl-w), causing hepatocyte apoptosis. Under MC-LR exposure, miR-122 agomir transfection could further increase the expressions of tumor suppressor genes and the release of cytochrome-c (Cyt-c) and decrease the expressions of Bcl-2 and Bcl-w. In conclusion, miR-122 reduction can mitigate MC-LR-induced apoptosis to a certain extent, which in turn, it is likely to have contributed to MC-LR-induced hepatocarcinogenesis.

Short-term effects of indoor and outdoor air pollution on the lung cancer morbidity in Henan Province, Central China.

Lung cancer is one of the most common cancer types and a major cause of death. The relationship between lung cancer morbidity and exposure to air pollutants is of particular concern. However, the relationship and difference in lung cancer morbidity between indoor and outdoor air pollution effects remain unclear. In this paper, the aim was to comprehensively investigate the spatial relationships between the lung cancer morbidity and indoor-outdoor air pollution in Henan based on the standard deviation ellipse, spatial autocorrelation analysis and GeoDetector. The results indicated that (1) the spatial distribution of lung cancer morbidity was related to the geomorphology, while high-morbidity areas were concentrated in the plains and basins of Central, Eastern and Southern Henan. (2) Among the selected outdoor air pollutants, PM2.5, NO2, SO2, O3 and CO were significantly correlated with the lung cancer morbidity. The degree of indoor air pollution was measured by the use of heating energy, and the proportions of coal-heating households, households with coal/biomass stoves and households with heated kangs were highly decisive in regard to the lung cancer morbidity. (3) The interaction between two factors was more notable than a single factor in explaining the lung cancer morbidity. Moreover, the interaction type was mainly nonlinear enhancement, and the proportion of households with coal/biomass stoves imposed the strongest interaction effect on the other factors.

Update on the adverse effects of microcystins on the liver.

Microcystins (MCs) are the most common cyanobacteria toxins in eutrophic water, which have strong hepatotoxicity. In the past decade, epidemiological and toxicological studies on liver damage caused by MCs have proliferated, and new mechanisms of hepatotoxicity induced by MCs have also been discovered and confirmed. However, there has not been a comprehensive and systematic review of these new findings. Therefore, this paper summarizes the latest advances in studies on the hepatotoxicity of MCs to reveal the effects and mechanisms of hepatotoxicity induced by MCs. Current epidemiological studies have confirmed that symptoms or signs of liver damage appear after human exposure to MCs, and a long time of exposure can even lead to liver cancer. Toxicological studies have shown that MCs can affect the expression of oncogenes by activating cell proliferation pathways such as MAPK and Akt, thereby promoting the occurrence and development of cancer. The latest evidence shows that epigenetic modifications may play an important role in MCs-induced liver cancer. MCs can cause damage to the liver by inducing hepatocyte death, mainly manifested as apoptosis and necrosis. The imbalance of liver metabolic homeostasis may be involved in hepatotoxicity induced by MCs. In addition, the combined toxicity of MCs and other toxins are also discussed in this article. This detailed information will be a valuable reference for further exploring of MCs-induced hepatotoxicity.

The latest advances in the reproductive toxicity of microcystin-LR.

Microcystin-LR (MC-LR) is an emerging environmental pollutant produced by cyanobacteria that poses a threat to wild life and human health. In recent years, the reproductive toxicity of MC-LR has gained widespread attention, a large number of toxicological studies have filled the gaps in past research and more molecular mechanisms have been elucidated. Hence, this paper reviews the latest research advances on MC-LR-induced reproductive toxicity. MC-LR can damage the structure and function of the testis, ovary, prostate, placenta and other organs of animals and then reduce their fertility. Meanwhile, MC-LR can also be transmitted through the placenta to the offspring causing trans-generational and developmental toxicity including death, malformation, growth retardation, and organ dysfunction in embryos and juveniles. The mechanisms of MC-LR-induced reproductive toxicity mainly include the inhibition of protein phosphatase 1/2 A (PP1/2 A) activity and the induction of oxidative stress. On the one hand, MC-LR triggers the hyperphosphorylation of certain proteins by inhibiting intracellular PP1/2 A activity, thereby activating multiple signaling pathways that cause inflammation and blood-testis barrier destruction, etc. On the other hand, MC-LR-induced oxidative stress can result in cell programmed death via the mitochondrial and endoplasmic reticulum pathways. It is worth noting that epigenetic modifications are also involved in reproductive cell apoptosis, which may be an important direction for future research. Furthermore, this paper proposes for the first time that MC-LR can produce estrogenic effects in animals as an environmental estrogen. New findings and suggestions in this review could be areas of interest for future research.

Microcystin-LR induces ovarian injury and apoptosis in mice via activating apoptosis signal-regulating kinase 1-mediated P38/JNK pathway.

As an emerging pollutant in the aquatic environment, microcystin-LR (MC-LR) can enter the body through multiple pathways, and then induce apoptosis and gonadal damage, affecting reproductive function. Previous studies focused on male reproductive toxicity induced by MC-LR neglecting its effects on females. The apoptotic signal-regulated kinase 1 (ASK1) is an upstream protein of P38/JNK pathway, closely associated with apoptosis and organ damage. However, the role of ASK1 in MC-LR-induced reproductive toxicity is unclear. Therefore, this study investigated the role of ASK1 in mouse ovarian injury and apoptosis induced by MC-LR. After MC-LR exposure, ASK1 expression in mouse ovarian granulosa cells was increased at the protein and mRNA levels, and decreased following pretreatment by antioxidant N-acetylcysteine, suggesting that MC-LR-induced oxidative stress has a regulatory role in ASK1 expression. Inhibition of ASK1 expression with siASK1 and NQDI-1 could effectively alleviate MC-LR-induced mitochondrial membrane potential damage and apoptosis in ovarian granulosa cells, as well as pathological damage, apoptosis and the decreased gonadal index in ovaries of C57BL/6 mice. Moreover, the P38/JNK pathway and downstream apoptosis-related proteins (P-P38, P-JNK, P-P53, Fas) and genes (MKK4, MKK3, Ddit3, Mef2c) were activated in vivo and vitro, but their activation was restrained after ASK1 inhibition. Data presented herein suggest that the ASK1-mediated P38/JNK pathway is involved in ovarian injury and apoptosis induced by MC-LR in mice. It is confirmed that ASK1 has an important role in MC-LR-induced ovarian injury, which provides new insights for preventing MCs-induced reproductive toxicity in females.

The activated ATM/p53 pathway promotes autophagy in response to oxidative stress-mediated DNA damage induced by Microcystin-LR in male germ cells.

Microcystin-LR (MC-LR) is an intracellular toxin with multi-organ toxicity and the testis is one of its important target organs. Although there is increasing research on MC-LR in male reproductive toxicity, the association between DNA damage and autophagy induced by MC-LR in male germ cells are still unclear. Therefore, it is important to explore the mechanism of MC-LR-induced DNA damage and the role of the activated ATM/p53 signaling pathway in testicular toxicity. The present study showed that MC-LR exposure significantly reduced gonadal index and induced pathological damage of the testes in mice. In addition, MC-LR increased the oxidative stress-related indicator hydroxyl radical, accompanied by increased levels of DNA damage-related indicators gamma-H2AX, 8-hydroxy-2'-deoxyguanosine, the olive tail moment (OTM) and DNA content of comet tail (TailDNA%) in trailing cells. Moreover, MC-LR activated the ATM/p53 pathway by enhancing the phosphorylation levels of ATM, CHK2 and p53 proteins, and then led to cell autophagy, ultimately triggering disrupted testicular cell arrangement, reduced sperm count and spermatogenic cell shedding. Importantly, after pretreatment with the antioxidant NAC, the expression levels of DNA damage-related indicators and the extent of damage in male germ cells were significantly reduced. Furthermore, pretreatment with the ATM inhibitor KU55933 could reduce the occurrence of autophagy and mitigate testicular toxicity of MC-LR through inhibiting the activation of the ATM/p53 pathway. These results indicate that MC-LR-induced oxidative stress can activate the DNA damage-mediated ATM/p53 signalling pathway to induce autophagy in male germ cells. This study provides a novel insight to further clarify the reproductive toxicity caused by MC-LR and to protect male reproductive health.

Design and engineering verification of an ultrashort throw ratio projection system with a freeform mirror.

A refractive-reflective combined ultrashort throw ratio projection optical system is designed. We use a freeform mirror to shorten the projection distance and correct distortion, and a plane mirror to turn back the optical path and reduce system volume. The projection system design method combines refractive lens group design and freeform surface mirror design with integrated optimization. The system's throw ratio is 0.11 with a projection distance of 320 mm and a 130 in. (1 in.=25.4 mm) screen size, which illustrates the advantages of the low throw ratio. The system's maximum distortion is 0.07%. To demonstrate the proposed system's performance, a prototype is developed. Experimental results confirm that the system performs excellently while meeting the design requirements. The system's advantages include low throw ratio, excellent imaging quality, miniaturization, and engineering feasibility.

A Phosphate-Starvation Induced RING-Type E3 Ligase Maintains Phosphate Homeostasis Partially Through OsSPX2 in Rice.

Phosphate (Pi), as the main form of phosphorus that can be absorbed by plants, is one of the most limiting macro-nutrients for plants. However, the mechanism for maintaining Pi homeostasis in rice (Oryza sativa) is still not well understood. We identified a Pi-starvation-induced E3 ligase (OsPIE1) in rice. Using an in vitro self-ubiquitination assay, we determined the E3 ligase activities of OsPIE1. Using GUS staining and GFP detection, we analyzed tissue expression patterns of OsPIE1 and the subcellular localization of its encoded protein. The function of OsPIE1 in Pi homeostasis was analyzed using OsPIE1 overexpressors and ospie1 mutants. OsPIE1 was localized to the nucleus, and expressed in epidermis, exodermis and sclerenchyma layers of primary root. Under Pi-sufficient condition, overexpression of OsPIE1 upregulated the expression of OsPT2, OsPT3, OsPT10 and OsPAP21b, resulting in Pi accumulation and acid phosphatases (APases) induction in roots. OsSPX2 was strongly suppressed in OsPIE1 overexpressors. Further comparative transcriptome analysis, tissue expression patterns and genetic interaction analysis indicated that the enhancing of Pi accumulation and APase activities upon overexpression of OsPIE1 was (at least in part) caused by repression of OsSPX2. These results indicate that OsPIE1 plays an important role in maintaining Pi homeostasis in rice.

Deletion of epithelial cell-specific Cdc42 leads to enamel hypermaturation in a conditional knockout mouse model.

Recent evidence suggests that GTPases Rho family plays an important role in tooth development; however, the role of Cdc42 in tooth development remains unclear. We aimed to investigate the function of Cdc42 in tooth development and amelogenesis. We generated an epithelial cell-specific K5-Cdc42 knockout (KO) mouse to evaluate post-eruption dental phenotypes using a K5-Cre driver line. This model overcomes the previously reported perinatal lethality. Tooth phenotypes were analyzed by micro X-ray, micro-computed tomography (CT), scanning electron microscopy (SEM), energy dispersive X-ray spectroscopy (EDX), wear rate, shear strength, and a microhardness test. Enamel matrix protein expression was determined by immunohistochemistry. KO mice displayed a hypomaturation phenotype, including incisors that lacked yellow pigmentation and were abnormally white, rapid attrition of molars following eruption, and decreased micro-hardness and shearing strength. Micro-CT data revealed that of incisor and molar enamel volumes were smaller in the KO than in wild-type (WT) mice. SEM analysis showed that the enamel prism structure was disordered. In addition, HE staining indicated a remarkable difference in the ameloblast morphology and function between KO and WT mice, and immunohistochemistry showed increased expression of amelogenin, ameloblastin, matrix metallopeptidase 20, kallikrein-related peptidase 4 and amelotin in the KO mice teeth. Our results suggest epithelium cell-specific Cdc42 deletion leads to tooth hypomaturation and transformation of the enamel prism structure that is likely due to altered ameloblast morphology and the secretion of enamel matrix proteins and proteases. This is the first in vivo evidence suggesting that Cdc42 is essential for proper tooth development and amelogenesis.

Mutation in SSUH2 Causes Autosomal-Dominant Dentin Dysplasia Type I.

Dentin dysplasia type I (DDI) is an autosomal-dominant genetic disorder resulting from dentin defects. The molecular basis of DDI remains unclear. DDI exhibits unique characteristics with phenotypes featuring obliteration of pulp chambers and diminutive root, thus providing a useful model for understanding the genetics of tooth formation. Using a large Chinese family with 14 DDI patients, we mapped the gene locus responsible for DDI to 3p26.1-3p24.3 and further identified a missense mutation, c.353C>A (p.P118Q) in the SSUH2 gene on 3p26.1, which co-segregated with DDI. We showed that SSUH2 (p.P118Q) perturbed the structure and significantly reduced levels of mutant (MT) protein and mRNA compared with wild-type SSUH2. Furthermore, MT P141Q knock-in mice (+/- and -/-) had a unique partial obliteration of the pulp cavity and upregulation or downregulation of six major genes involved in odontogenesis: Dspp, Dmp1, Runx2, Pax9, Bmp2, and Dlx2. The phenotype of missing teeth was determined in zebrafish with morpholino gene knockdowns and rescued by injection of normal human mRNA. Taken together, our observations demonstrate that SSUH2 disrupts dental formation and that this novel gene, together with other odontogenesis genes, is involved in tooth development.

RhoA regulates Activin B-induced stress fiber formation and migration of bone marrow-derived mesenchymal stromal cell through distinct signaling.

BACKGROUND: In our previous study, Activin B induced actin stress fiber formation and cell migration in Bone marrow-derived mesenchymal stem cells (BMSCs) in vitro. However, the underlying molecular mechanisms are not well studied. RhoA is recognized to play a critical role in the regulation of actomyosin cytoskeletal organization and cell migration. METHODS: Pull-down assay was performed to investigate the activity of RhoA. The dominant-negative mutants of RhoA (RhoA(N19)) was used to determine whether RhoA has a role in Activin B-induced cytoskeleton organization and cell migration in BMSCs. Cytoskeleton organization was examined by fluorescence Rhodamine-phalloidin staining, and cell migration by transwell and cell scratching assay. Western blot was carried out to investigate downstream signaling cascade of RhoA. Inhibitor and siRNAs were used to detect the role of downstream signaling in stress fiber formation and/or cell migration. RESULTS: RhoA was activated by Activin B in BMSCs. RhoA(N19) blocked Activin B-induced stress fiber formation and cell migration. ROCK inhibitor blocked Activin B-induced stress fiber formation but enhanced BMSCs migration. Activin B induced phosphorylation of LIMK2 and Cofilin, which was abolished by ROCK inhibition. Both of siRNA LIMK2 and siRNA Cofilin inhibited Activin B-induced stress fiber formation. CONCLUSIONS: RhoA regulates Activin B-induced stress fiber formation and migration of BMSCs. A RhoA-ROCK-LIMK2-Cofilin signaling node exists and regulates actin stress fiber formation. RhoA regulates Activin B-induced cell migration independent of ROCK. GENERAL SIGNIFICANCE: Better understanding of the molecular mechanisms of BMSCs migration will help optimize therapeutic strategy to target BMSCs at injured tissues.

A splicing mutation in VPS4B causes dentin dysplasia I.

BACKGROUND: Dentin dysplasia I (DDI) is a genetically heterogeneous autosomal-dominant disorder characterised by rootless teeth with abnormal pulpal morphology, the aetiology of which presents as genetically heterogeneous. METHODS AND RESULTS: Using a cohort of a large Chinese family with 10 patients with DDI, we mapped to a 9.63 Mb candidate region for DDI on chromosome 18q21.2-q21.33. We then identified a mutation IVS7+46C>G which resulted in a novel donor splice site in intron 7 of the VPS4B gene with co-segregation of all 10 affected individuals in this family. The aberrant transcripts encompassing a new insert of 45 bp in size were detected in gingival cells from affected individuals. Protein structure prediction showed that a 15-amino acid insertion altered the ATP-binding cassette of VPS4B. The mutation resulted in significantly reduced expression of mRNA and protein and altered subcellular localisation of VPS4B, indicating a loss of function of VPS4B. Using human gingival fibroblasts, the VPS4B gene was found to act as an upstream transducer linked to Wnt/ß-catenin signalling and regulating odontogenesis. Furthermore, knockdown of vps4b in zebrafish recapitulated the reduction of tooth size and absence of teeth similar to the tooth phenotype exhibited in DDI index cases, and the zebrafish mutant phenotype could be partially rescued by wild-type human VPS4B mRNA. We also observed that vps4b depletion in the zebrafish negatively regulates the expression of some major genes involved in odontogenesis. CONCLUSIONS: This study identifies VPS4B as a disease-causing gene for DDI, which is one of the important contributors to tooth formation, through the Wnt/ß-catenin signalling pathway.

Beneficial Effects of Fecal Microbiota Transplantation on Ulcerative Colitis in Mice.

BACKGROUND: Ulcerative colitis (UC) is a chronic condition and the most common form of inflammatory bowel disease. The goal of standard treatment is mainly to induce and maintain remission with anti-inflammatory, immunosuppressive agents, and/or colectomy. Fecal microbiota transplantation (FMT) has been used successfully to treat relapsing or refractory Clostridium difficile infection. The alteration of microbiota in mouse models of UC as well as in patients suggested the possibility of treating UC with FMT. AIMS: To study the effects of FMT on dextran sodium sulfate (DSS)-induced UC model in mice. METHODS: Littermates of BALB/c and C57BL/6J were randomized into four groups: normal control , treatment with DSS for 7 days (DSS - FMT), treatment with DSS followed by FMT for another 8 days (DSS + FMT), and treatment with DSS and FMT followed by another 5 days for recovery (remission). Body weight, survival rate, and DAI scores of mice in each group were recorded. Changes in distal colon were studied by histopathology. Alterations of spleen and lamina propria regulatory lymphocytes, major bacterial species in feces and inflammatory cytokines in colon were also studied. RESULTS: C57BL/6J mice experienced more significant weight loss than BALB/c mice after DSS treatment, regardless of whether the two strains of mice were co-housed or not. FMT caused reversal of DAI scores in BALB/c but not in C57BL/6J mice. In BALB/c mice, FMT also reduced colon inflammation that was paralleled by decreased inflammatory cytokine levels, altered bacterial microbiota, and regulatory lymphocyte proportions. CONCLUSIONS: FMT is effective in a mouse model of UC through its modulation on gut microbiota and the host immune system.

[Screening of CTSC gene mutations in a Chinese pedigree affected with Papillon-Lefevre syndrome].

OBJECTIVE: To analyze the clinical phenotype of a Chinese pedigree affected with Papillon-Lefevre syndrome(PLS) and detect mutation of CTSC gene. METHODS: Clinical phenotypes were noted, and oral examination for the proband was carried out for the clinical diagnosis of PLS. PCR and Sanger sequencing were used to identify potential mutation of the CTSC gene. Functional effect of the mutation was predicted with SIFT and PolyPhen-2. Swiss-Port was used to predict the tertiary structure of wild type and mutant proteins. The mRNA and protein expression were analyzed by real-time PCR and Western blotting. RESULTS: A homozygous mutation c.901G>A (p.G301S) in exon 7 of CTSC gene was identified in the patient. Both parents of the patient had carried a heterozygous c.901G>A mutation. The mutation was located in the conserved region of CTSC enzyme and was predicted to be damaging by changing the structure of the protein, which could affect the activity of Cathepsin C. However, no significant difference was found in the expression of p.G301S variant at the mRNA and protein levels compared with that of the wild type CTSC gene. CONCLUSION: The c.901G>A mutation of the CTSC gene was first reported in China, which has expanded its mutation spectrum.