OsMGD1-Mediated Membrane Lipid Remodeling Improves Salt Tolerance in Rice.

Salt stress severely reduces photosynthetic efficiency, resulting in adverse effects on crop growth and yield production. Two key thylakoid membrane lipid components, monogalactosyldiacylglycerol (MGDG) and digalactosyldiacylglycerol (DGDG), were perturbed under salt stress. MGDG synthase 1 (MGD1) is one of the key enzymes for the synthesis of these galactolipids. To investigate the function of OsMGD1 in response to salt stress, the OsMGD1 overexpression (OE) and RNA interference (Ri) rice lines, and a wild type (WT), were used. Compared with WT, the OE lines showed higher chlorophyll content and biomass under salt stress. Besides this, the OE plants showed improved photosynthetic performance, including light absorption, energy transfer, and carbon fixation. Notably, the net photosynthetic rate and effective quantum yield of photosystem II in the OE lines increased by 27.5% and 25.8%, respectively, compared to the WT. Further analysis showed that the overexpression of OsMGD1 alleviated the negative effects of salt stress on photosynthetic membranes and oxidative defense by adjusting membrane lipid composition and fatty acid levels. In summary, OsMGD1-mediated membrane lipid remodeling enhanced salt tolerance in rice by maintaining membrane stability and optimizing photosynthetic efficiency.

Radiomic features from multiparametric magnetic resonance imaging predict molecular subgroups of pediatric low-grade gliomas.

BACKGROUND: We aimed to develop machine learning models for prediction of molecular subgroups (low-risk group and intermediate/high-risk group) and molecular marker (KIAA1549-BRAF fusion) of pediatric low-grade gliomas (PLGGs) based on radiomic features extracted from multiparametric MRI. METHODS: 61 patients with PLGGs were included in this retrospective study, which were divided into a training set and an internal validation set at a ratio of 2:1 based on the molecular subgroups or the molecular marker. The patients were classified into low-risk and intermediate/high-risk groups, BRAF fusion positive and negative groups, respectively. We extracted 5929 radiomic features from multiparametric MRI. Thereafter, we removed redundant features, trained random forest models on the training set for predicting the molecular subgroups or the molecular marker, and validated their performance on the internal validation set. The performance of the prediction model was verified by 3-fold cross-validation. RESULTS: We constructed the classification model differentiating low-risk PLGGs from intermediate/high-risk PLGGs using 4 relevant features, with an AUC of 0.833 and an accuracy of 76.2% in the internal validation set. In the prediction model for predicting KIAA1549-BRAF fusion using 4 relevant features, an AUC of 0.818 and an accuracy of 81.0% were achieved in the internal validation set. CONCLUSIONS: The current study demonstrates that MRI radiomics is able to predict molecular subgroups of PLGGs and KIAA1549-BRAF fusion with satisfying sensitivity. TRIAL REGISTRATION: This study was retrospectively registered at clinicaltrials.gov (NCT04217018).

Antibiotic resistance genes and bacterial community dynamics in the seawater environment of Dapeng Cove, South China.

In recent years, the propagation of antibiotic resistance genes (ARGs) and increased antibiotic resistance in pathogens have gained serious attention. Numerous reports have investigated the influence of domestic sewage discharge, medical wastewater and aquaculture wastewater on rivers and lakes, while the dynamics of ARGs in seawater and the relationships between ARGs, bacterial community structure and environmental factors have been less thoroughly described. In this study, the abundance, distribution and source of ARGs, as well as the relationships between ARGs, bacterial community changes and environmental factors in the seawater environment and sediment of Dapeng Cove, were investigated. Real-time quantitative PCR and Illumina Miseq sequencing technology were applied to determine the effects of the production cycle of cage culture, tourism and seasonality on ARGs. Chloramphenicol resistance genes (floR, cmlA) and sulfonamide resistance genes (sul1) were the dominant resistance genes in water and sediment. Pearson's correlation analysis showed that the abundance of all ARGs and the integrase I gene intI1 was positively correlated with chemical oxygen demand and suspended solids. Class 1 integrons might facilitate the dissemination of ARGs, and intI1 was detected in all samples at high concentrations. In aqueous environments, Cyanobacteria, Proteobacteria and Bacteroidetes were the dominant phyla, among which Proteobacteria and Bacteroidetes were positively correlated with the concentration of target ARGs. In the sediment, Proteobacteria, Bacteroidetes, Chloroflexi, Acidobacteria and Planctomycetes were the dominant phyla, among which Bacteroidetes and Planctomycetes were positively correlated with most of the target ARGs and had a significant influence on changes in the abundance of ARGs. The domestic sewage was the main source of ARGs in the seawater. Our results showed that bacterial community structure and environmental factors affected the distributional dynamics of ARGs. Anthropogenic activities played significant roles in promoting ARGs abundance in the seawater environments.

Contamination of antibiotic resistance genes (ARGs) in a typical marine aquaculture farm: source tracking of ARGs in reared aquatic organisms.

Although the prevalence and concentrations of antibiotic resistance genes (ARGs) in aquaculture is receiving increasing scientific interest, there is little understanding of the direct sources and dissemination pathways of ARGs in marine aquaculture-reared organisms. This study investigated the dynamics of ARGs and the bacterial community throughout the rearing period in a typical marine aquaculture farm in South China. The results demonstrated that sul1 and qnrD were predominant in the sediment, and qnrD and qnrA were predominant in the intestinal tracts of shrimps. Network analysis showed that the chemical oxygen demand, total organic carbon, dissolved organic carbon, suspended solids, and total phosphorus were positively correlated with the predominant ARGs. The results of the network and source tracking analyses indicate that environmental factors and the bacterial community may drive the dissemination of ARGs dissemination in the environment and in shrimp reared by marine aquaculture, and sediment is the most direct and important medium in this dissemination. These results aid in improving our understanding of the sources, level, and dissemination of ARGs in marine aquaculture.

Abundance and removal of antibiotic resistance genes (ARGs) in the rearing environments of intensive shrimp aquaculture in South China.

Although research regarding antibiotic resistance genes (ARGs) in aquaculture environments has gained increasing scientific interest, further studies are required to understand the abundances and removal mechanisms of ARGs during the entire rearing period of shrimp aquaculture. Thus, in this study, abundances, distributions and removal rates of ARGs in different environmental compartments of intensive shrimp farms in South China were investigated during the entire rearing period. The results indicated that sul1 and cmlA were the predominant ARGs in the water and sediment samples. Additionally, the total abundance of ARGs was higher in shrimp pond water than in the source water and farm effluent. Moreover, sediment samples indicated significantly higher ARG abundances than water samples from the shrimp ponds (P < 0.05). Environmental factors were found to significantly affect the distribution of ARGs in shrimp rearing environments. Furthermore, stable ponds aided the removal of ARGs from shrimp pond water. This study accounted for temporal variations in ARG abundances as well as removal of ARGs in different environmental compartments during the entire shrimp rearing period. However, additional research is required to optimize the water treatment process for removal of ARGs from the aquaculture.

[Murine model of busulfan-induced spermatogenesis regeneration: a quantitative evaluation].

OBJECTIVE: To quantitatively evaluate the murine model of spermatogenesis regeneration induced by two-dose busulfan injection. METHODS: Fifty-four male mice were randomly divided into a control and two model groups of equal number, the former treated by two-dose intraperitoneal injection of 50% DMSO solution at 10 ml/kg, and the latter by that of busulfan at 10 mg/kg and 15 mg/kg respectively to establish spermatogenesis regeneration models, both at the interval of 24 days between the two doses. Spermatogenesis in seminiferous epithelia was evaluated by Johnsen score, and the expressions of GATA-4 and GDNF mRNA in Sertoli cells were detected by real time quantitative PCR at 3, 4 and 8 weeks after the treatment. RESULTS: Johnsen score kept stable in the control group at all stages (P > 0.05), but higher than in the model groups at 3 and 4 weeks (P < 0.01). It was lower in the 15 mg/kg than in the 10 mg/kg model group at 4 and 8 weeks (P < 0.01) , and than in the control group at 8 weeks (P < 0.05), but had no significant difference between the 10 mg/kg and the control groups (P > 0.05). Nor did the expression of GATA-4 mRNA in Sertoli cells show any significant difference among the three groups at different stages after the treatment (P > 0.05), and that of GDNF mRNA at different stages in the control group (P > 0.05). Compared with the controls, the level of GDNF mRNA in Sertoli cells was significantly higher at 3 weeks but lower at 4 weeks in the model groups (P < 0.01), and lower in the 15 mg/kg group (P < 0.01) and comparable in the 10 mg/kg group at 8 weeks (P > 0.05); and it was lower in the 15 mg/kg than in the 10 mg/kg group at all stages (P < 0.01). CONCLUSION: Two-dose intraperitoneal injection of 10 mg/kg busulfan at the interval of 24 days is an optimal option for the establishment of a murine model of spermatogenesis regeneration. Higher dose of busulfan may induce deficient expression of GDNF in Sertoli cells and result in incomplete restoration of spermatogenesis.

Long-term culture and transplantation of spermatogonial stem cells from BALB/c mice.

Development of a culture system that supports self-renewal and proliferation of spermatogonial stem cells (SSCs) is enormously valuable for experimental research and potential treatment for male infertility. Although several research groups had reported their successes in SSC isolation and culture, the two current accepted culture systems are different in cell enrichment methods, serum and growth factors. Previous researches also indicated SSCs from different mouse strains required different culture conditions. Here we report for the first time that SSCs from BALB/c mice could be cultured in an improved culture system for 3 months. The modified culture system consisted of an improved enzymatic procedure, the enrichment of undifferentiated spermatogonia by differential adherence selection of isolated SSCs, mouse embryonic fibroblast feeder cells, StemPro-34 SFM medium supplemented with glial cell line-derived neurotrophic factor (GDNF), basic fibroblast growth factor and GDNF-family receptor alpha1 (GFRalpha1). The improved digestion method increased the viability and enrichment efficiency of isolated testis cells. Furthermore, basal culture medium with 10% fetal bovine serum as selected medium could increase the number of germ cell colonies in the initiation stage of culture. Cultured SSCs were characterized morphologically and formed typical colonies. Immunocytochemical staining and RT-PCR showed that cultured SSCs expressed Oct-4, GFRalpha1, Sox2 and several other special genes resembling undifferentiated spermatogonia. Spermatogonia transplantation further confirmed that cultured SSCs were functionally normal and could restore complete spermatogenesis. The culture methods described here could serve as a paradigm to establish conditions for the culture of SSCs from other species, allowing identification of universal factors necessary for proliferation of SSCs.

[Reconstruction of rabbit urethra using urethral extracellular matrix].

OBJECTIVE: To evaluate the feasibility of reconstruction of rabbit urethra using urethral extracellular matrix. METHODS: Extracellular matrix was obtained from the urethra of 20 donor New Zealand rabbits. In experimental group, 20 rabbits underwent segmental urethral resection (about 1.0 to 1.5 cm in length) and the defects were replaced by a tube of extracellular matrix. The serum TNFalpha was detected by ELISA to assess the immunity response preoperatively and 12, 24, 48 h postoperatively. The regenerated urethral segments were taken for histologic and pathologic study 10 days, 3 weeks, 6 weeks and 24 weeks after operation. The urodynamics, urethroscopy and urethrography were also performed. RESULTS: The serum TNFalpha in experiment group slightly rised, with no significant difference when compared with that in control group. 10 days after operation, epithelial cell migrated into the extracellular matrix from two ends, and small vessels were also found. 3 weeks later, several layers of urothelium covered the whole surface of the matrix tube. 6 weeks later, the irregularly arranged smooth muscle fibers were fist observed by Van Gieson staining. 24 weeks after operation, the smooth muscle cells increased, the appearance of the regenerated urethra segments were very similar to normal urethral wall components. The urethrography and urodynamic evaluation revealed no difference between the normal and the regenerated urethral tube. CONCLUSIONS: The urethral extracellular matrix might be an ideal replacement material for urethral defect.

[Long-term culture and identification of spermatogonial stem cells from BALB/c mice in vitro].

OBJECTIVE: To establish a long-term culture system for mouse spermatogonial stem cells (SSCs) and to discuss the key factor that supports mouse SSC self-renewal and proliferation. METHODS: Testis cells from 4-6 days postpartum male transgenic BALB/c mce were collected by a modified two-step enzymatic digestion method and plated on 0. 2% elatin-coated tissue culture plates. The germ cells were enriched by differential adherence selections after respectively incubated for 1, 5 and 24 h and then plated on the mitomycin C-inactivated mouse embryonic fibroblast (MEF) feeder layer. The basal culture medium was StemPro-34 SFM supplemented with other 15 nutrient factors. The 20 ng/ml Glial cell line-derived neurotrophic factor (GDNF), 10 ng/ml basic fibroblast growth factor (bFGF) and 200 ng/ml GDNF-family receptor alpha 1 (GFRalpha1) were added to the serum-free medium to promote SSC proliferation. Several important surface markers and special genes were examined by immunocytochemical staining and RT-PCR analysis. RESULTS: After 3-4 days culture on the MEF feeder, SSCs proliferated continuously and formed typical colonies. SSCs from the BALB/c mice could be cultured in a steady state for 3 months. Immunocytochemical staining showed that Oct4 was specifically expressed in the cultured SSC nucleus and GFRalpha1 strongly expressed on the surface of the membrane. RT-PCR confirmed that the cultured SSCs expressed Oct-4, GFRalpha1, Sox2 and several other special genes resembling undifferentiated spermatogonia. CONCLUSION: SSCs from BALB/c mice could be cultured in the improved culture system for 3 months. This culture system could help further understand the regulating mechanism of SSCs and might provide an opportunity for the treatment of male infertility by SSC transplantation.

Curative effect and histocompatibility evaluation of reconstruction of traumatic defect of rabbit urethra using extracellular matrix.

OBJECTIVE: To investigate the curative effect and histocompatibility of reconstruction of traumatic urethral defect of rabbit using urethral extracellular matrix (ECM). METHODS: Urethral ECM was obtained by excision of the urethra in 20 donor rabbits. In experimental group, 20 rabbits were resected a 1.0 cm-1.5 cm segment of the urethra and artificially made a model of traumatic urethral defect, then reconstructed by the urethral extracellular matrix of the same length. The rabbit immunity response was assessed by lymphocyte transformation test and serum TNF-alpha level. The reconstructed urethral segments were stained with hematoxylin-eosin and Van Gieson stain and observed by histological examination postoperatively. The urethrography, urethroscopy and urodynamic examinations were performed. RESULTS: There was no significant difference in stimulative index of lymphocyte transformation between ECM group and control group. The serum TNF-alpha levels of ECM group slightly rose, but the increase was not significant as compared with control group. On postoperative day 10, epithelial cell had migrated from each side and small vessels were found in the extracellular matrix. In the 3rd week, several layers of urothelium covered the whole surface of the matrix tube. In the 6th week, the disorganized arrangements of smooth muscle fibers were firstly observed by Van Gieson staining. In the 24th week, the smooth muscle cells increased and the matrix tube appeared fairly similar to normal urethral wall components. The urethroscopy and urodynamic evaluation revealed that the surface of reconstructed urethra was smooth and emiction was unobstructed. CONCLUSION: The urethral extracellular matrix might be an ideal and safe biomaterial for the reconstruction of urethral traumatic defect.

[Sorting of spermatogonial stem cells in mice].

OBJECTIVE: To search for an effective method for enriching spermatogonial stem cells in mice. METHODS: Bilateral artificial cryptorchidism was performed on 20 six-week old male Kunming mice. Three months after the operation, the testes were removed and single cell suspension prepared by two-step enzyme digestion. FITC-conjugated anti-alpha6-integrin antibody and PE-conjugated anti-c-kit antibody were added for adequate time on ice. Then the cells with low side scatter light-scattering properties were sorted and positively stained for alpha6-integrin and negative c-kit expression. And the viability of the isolated cells was assessed by trypan blue exclusion. RESULTS: The sorted spermatogonial stem cells constitute 2.8% of the testis cells and over 95% of them were viable. CONCLUSION: FACS can be used to isolate quantities of viable spermatogonial stem cells.

[Treatment of buried penis in children with fixation at the dartos coat of penis through prepubic route].

OBJECTIVE: To observe the long-time results of the treatment of buried penis with fixation at the base of penis through pre-pubic route. METHODS: From Aug 2002 to Dec 2003, the procedure was performed in 34 children without penile skin insufficiency. The major technique involved the release of hided penis shaft by partly shearing suspensory ligament, fixating the bucks fascia and tunica albuginea at 3 and 9 o'clock positions with the subcutaneous penile skin at the base of penis just under the pubic symphysis. RESULTS: Twenty-one patients have been reviewed over one year with satisfactory appearance. Thirteen patients were lost to follow-up. No recurrence happened in our group. CONCLUSION: We compare this technique with major treatment methods for buried penis in children at present, and we conclude this technique is a reasonable, simplified and aesthetical method for the treatment of buried penis in children.

[Modified technique of testicular autotransplantation in dogs].

OBJECTIVE: To establish dog model of testicular autotransplantation with a modified technique. METHODS: Testicular autotransplantations were performed on the right side of 30 male dogs, whose ages ranged from 1.5 to 2.0 years old and weights ranged from 14 to 17 kg. After the spermatic artery with a cuff of abdominal aorta and spermatic vein and with a cuff of inferior vena cava were detached, the testis was perfused and kept at icing temperature. An end-to-side anastomosis of the spermatic vessels to the external iliac vessels was conducted subsequently. The survival conditions of the autografts were assessed by digital subtraction arteriography (DSA). Histological examination and detection on the serum levels of follicle stimulating hormone (FSH), luteotrophic hormone (LH), and testosterone (T) were made at two weeks intervals. RESULTS: Of the 30 testicular autotransplantations performed, 27 cases were successful. The success rate was 90%. The time of heat ischemia, cold ischemia, anastomosis of spermatic vessels, and total operation was 4.5+/-0.9 minutes, 50.0+/-10.0 minutes, 35.5+/-5.5 minutes, and 3.5+/-0.5 hours respectively. DSA proved that the testis survived well. No morphological abnormality was found at different stages of the spermatogenic cells. The LH level was higher than that before operation, being statistically different (P<0.05); however, the levels of FSH and T did not changed significantly (P>0.05). CONCLUSION: A stable and feasible model of testicular autotransplantation is established and it provides a reliable experimental platform for human testicular transplantation.

[Modified technique of testis autotransplantation in dogs].

OBJECTIVE: To establish a dog model of testis autotransplantation with a modified technique. METHODS: Testis autotransplantations were performed in 30 dogs. After detachment of the spermatic artery with a cuff of the abdominal aorta and the spermatic vein with a cuff of inferior vena cava, the testis was perfused and kept at ice temperature. An end-to-side anastomosis of the spermatic vessels to the external iliac vessels was conducted. RESULTS: The success rate of the testis autotransplantations was 90% (27/30) and the time for heat ischemia, cold ischemia, anastomosis of spermatic vessels and the whole operation were (4.5 +/- 0.9) minutes, (50.0 +/- 5.0) minutes, (35.5 +/- 5.5) minutes and (3.5 +/- 0.5) hours respectively. CONCLUSION: A stable and feasible model of testis autotransplantation was established, which provides a reliable experimental base for testis autotransplantation.

[Advances in testis transplantation].

Testis transplantation is an important and effective way to treat abdominal impalpable cryptorchidism, male hypogonadism and male infertility. Since 1990s a lot of advances have been made in microsurgical autotransplantation, homotransplantation, testicular tissue transplantation and Leydig cell transplantation. The main achievements include the application of laparoscopy in autotransplantation, researches on the influential factors in spermatogenesis after homotransplantation, explorations of new treatment methods such as fetal testis transplantation, spermatogonial stem cell transplantation and so on. The advances in testis transplantation are summarized in this paper based on the related literature of recent years.

Reconstruction of rabbit urethra using urethral extracellular matrix.

BACKGROUND: Urethral reconstruction for both congenital and acquired etiologies remains a challenge for most urologic surgeons. Tissue engineering has been proposed as a strategy for urethral reconstruction. The purpose of this study was to determine whether a naturally derived extracellular matrix substitute developed for urethral reconstruction would be suitable for urethral repair in an animal model. METHODS: A urethral segmental defect was created in 20 male rabbits. The urethral extracellular matrix, obtained and processed from rabbit urethral tissue, was trimmed and transplanted to repair the urethral defect. Then, the regenerated segment was studied histologically by haematoxylin-eosin staining and Van Gieson staining at 10 days, 3 weeks, 6 weeks, and 24 weeks postoperation. Retrograde urethrography was used to evaluate the function of the regenerated urethras of 4 rabbits 10 and 24 weeks after the operation. The urodynamics of 4 rabbits from the experimental group and control group I were assessed and compared. In addition, 4 experimental group rabbits were examined by a urethroscope 24 weeks after the operation. RESULTS: At 10 days after operation, epithelial cells had migrated from each side, and small vessels were observed in the extracellular matrix. The matrix and adjacent areas of the host tissue were infiltrated with inflammatory cells. The epithelium covered the extracellular matrix fully at 3 weeks postoperation. Well-formed smooth-muscle cells were first confirmed after 6 weeks, at which point the inflammatory cells had disappeared. At 24 weeks postoperation, the regenerated tissue was equivalent to the normal urethra. Urethrography and urodynamic evaluations showed that there was no difference between normal tissue and regenerated tissue. CONCLUSIONS: Urethral extracellular matrix appears to be a useful material for urethral repair in rabbits. The matrix can be processed easily and has good characteristics for tissue handling and urethral function.

[Effects of icariin on the erectile function and expression of nitrogen oxide synthase isoforms in corpus cavernosum of arterigenic erectile dysfunction rat model].

OBJECTIVE: The study the effects of oral administered icariin on intracavernosal pressure (ICP) and on expression of the nitrogen oxide synthase (NOS) isoforms in corpus cavernosum (CC) of arteriogenic erectile dysfunction (A-ED) rat model. METHODS: Forty adult male Wistar rats were randomly divided into 4 groups of 10 rats: shame operated group (group A) and three A-ED model groups (group B, C and D). The internal pudendal arteries were isolated and ligated with 7-O nylon thread at both the main trunk and the penile branches to establish the A-ED model. ICP were tested after the operation to make sure the successful model establishment. The groups A and B were treated with saline: and the groups C and D were treated with icariin (5 mg/kg/day and 10mg/kg/day respectively) orally for 30 days. Then the ICP was measured again. The tissues of corpus cavernosum were taken and RT-PCR was used to detect the mRNA expression of nNOS, iNOS and eNOS in CC, and Western-blot was used to detect the protein expression of these NOS isoforms. RESULTS: The ICP in the group B was significantly decreased compared to the group A (P < 0.01), but the ICP values in the groups C and D were both increased compared to those in the group B (both P < 0.01). The expressions of the mRNA and protein of nNOS, iNOS, and eNOS were all decreased in the group B, however, the mRNA and protein expressions of eNOS were increased a in the groups C and D. In the group C, iNOS also increased. The expression of nNOS showed no obvious changes in the group C and group D. CONCLUSION: Chronic oral treatment with Icariin increases the erectile function (ICP) and restores the eNOS expression in CC of A-ED rats. Icariin may have a long-term therapeutic effect on ischemia/hypoxia induced ED.

[Extracellular matrix for the replacement of ureteral defect].

This is an experiment on rabbits to evaluate the possibility of ureteral replacement by extracellular matrix. We adopted a biochemical method for preparing a new tissue engineering material named Extracellular Matrix (ECM), and the ECMs were used as homologous grafts to replace the defect in the ureters. Light microscopy, scanning electron microscopy, immunohistochemical technique and intravenous urography were used. The routine blood and biochemical laboratory tests were made before and after operation, and the measured values of pressure in the ureter of experiment and control groups were compared. The ureteral ECM was found in the experiment to promote the regeneration of all ureteral wall components. There were no significant differentces between the regenerative tissue and the normal tissue in morphology and function 16 weeks after replacement. The homologous ECM might be an ideal replacement material for ureteral defect.