Prenatal prednisone exposure disturbs fetal kidney development and its characteristics.

Prednisone is a synthetic glucocorticoid that is commonly used in both human and veterinary medication. Now, it is also recognized as an emerging environmental contaminant. Pregnant women may be exposed to prednisone actively or passively through multiple pathways and cause developmental toxicity to the fetus. However, the impact of prenatal prednisone exposure (PPE) on fetal kidney development remains unclear. In this study, pregnant mice were administered prednisone intragastrically during full-term pregnancy with different doses (0.25, 0.5, or 1 mg/(kg·day)), or at the dose of 1 mg/(kg·day) in different gestational days (GD) (GD0-9, GD10-18, or GD0-18). The pregnant mice were euthanized on GD18. HE staining revealed fetal kidney dysplasia, with an enlarged glomerular Bowman's capsule space and a reduced capillary network in the PPE groups. The expression of the podocyte and the mesangial cell marker genes was significantly reduced in the PPE groups. However, overall gene expression in renal tubules and collecting ducts were markedly increased. All of the above effects were more pronounced in high-dose, full-term pregnancy, and female fetuses. Studies on the mechanism of the female fetal kidney have revealed that PPE reduced the expression of Six2, increased the expression of Hnf1ß, Hnf4α, and Wnt9b, and inhibited the expression of glial cell line-derived neurotrophic factor (GDNF) and Notch signaling pathways. In conclusion, this study demonstrated that there is a sex difference in the developmental toxicity of PPE to the fetal kidney, and the time effect is manifested as full-term pregnancy > early pregnancy > mid-late pregnancy.

Low H3K27 acetylation of SF1 in PBMC: a biomarker for prenatal dexamethasone exposure-caused adrenal insufficiency of steroid synthesis in male offspring.

Dexamethasone is widely used to treat pregnancy disorders related to premature delivery. However, lots of researches have confirmed that prenatal dexamethasone exposure (PDE) could increase the risk of offspring multiple diseases. This study was designed to elucidate the epigenetic mechanism of adrenal developmental programming and explore its early warning marker in peripheral blood mononuclear cells (PBMC). We found the adrenal morphological and functional changes of PDE male offspring rats before and after birth, which were mainly performed as the decreased serum corticosterone concentration, steroidogenic acute regulatory (StAR) protein expression, and histone 3 lysine 27 acetylation (H3K27ac) level of steroidogenic factor 1 (SF1) promoter region and its expression. Simultaneously, the expressions of glucocorticoid receptor (GR) and histone acetylation enzyme 5 (HDAC5) in the PDE male fetal rats were increased. In vitro, dexamethasone reduced the expression of SF1, StAR, and cortisol production and still increased the expression of GR and HDAC5, the binding between GR and SF1 promoter region, and protein interaction between GR and HDAC5. GR siRNA or HDAC5 siRNA was able to reverse the above roles of dexamethasone. Furthermore, in vivo, we confirmed that H3K27ac levels of SF1 promoter region and its expression in PBMC of the PDE group were decreased before and after birth, showing a positive correlation with the same indexes in adrenal. Meanwhile, in clinical trials, we confirmed that prenatal dexamethasone application decreased H3K27ac of SF1 promoter region and its expression in neonatal PBMC. In conclusion, PDE-caused adrenal insufficiency of male offspring rats was related to adrenal GR activated by dexamethasone in uterus. The activated GR, on the one hand, increased its direct binding to SF1 promoter region to inhibit its expression, on the other hand, upregulated and recruited HDAC5 to decrease H3K27ac level of SF1 promoter region, and strengthened the inhibition of SF1 and subsequent StAR expression.

Balance training benefits chronic ankle instability with generalized joint hypermobility: a prospective cohort study.

BACKGROUND: Balance training is the first choice of treatment for chronic ankle instability (CAI). However, there is a lack of research on the effects of balance training in CAI with generalized joint hypermobility (GJH). This study is to compare the outcomes of balance training in CAI patients with and without GJH. METHODS: Forty CAI patients were assigned into the GJH group (Beighton ≥ 4, 20) and non-GJH group (Beighton < 4, 20) and they received same 3-month supervised balance training. Repeated measure ANOVA and independent t test were used to analyze self-reported questionnaires (Foot and ankle ability measure, FAAM), the number of patients experiencing ankle sprain, isokinetic muscle strength and postural control tests (Star excursion balance test, SEBT and Balance errors system, BES) before training, post-training immediately, and post-training 3 months, respectively. RESULTS: At baseline, no differences were found between groups with except for GJH group having poorer SEBT in the posteromedial direction (83.6 ± 10.1 vs 92.8 ± 12.3, %) and in the posterolateral direction (84.7 ± 11.7 vs 95.7 ± 8.7, %). Following the balance training, GJH group demonstrated lower re-sprain ratio (immediately after training, 11.1% vs 23.5%, 3 month after training, 16.7% vs 29.4%) than non-GJH group, as well as greater FAAM-S score, plantarflexion strength and dorsiflexion strength at post-training immediately and 3 months, and both groups improved similarly in the FAAM-A score, muscle strength and balance control (SEBT in the posterior-lateral and posterior-medial directions, and BES scores) compared with baseline. CONCLUSIONS: CAI patients with GJH gained equally even better postural stability and muscle strength after the balance training than the non-GJH patients. Balance training could still be an effective treatment for CAI patients with GJH before considering surgery. TRIAL REGISTRATION: ChiCTR1900023999, June 21st, 2019.

Arthroscopic Anatomical Double-Bundle Medial Patellofemoral Complex Reconstruction Improves Clinical Outcomes in Treating Recurrent Patellar Dislocation Despite Trochlear Dysplasia, Elevated Tibial Tubercle-Trochlear Groove Distance, and Patellar Alta.

PURPOSE: To evaluate the clinical outcomes of arthroscopically assisted double-bundle medial patellofemoral complex reconstruction (MPFC-R). METHODS: A retrospective review was carried out among adult patients who experienced at least 2 patellar dislocations and underwent primary arthroscopically assisted MPFC-R between January 2014 and November 2019. Dejour classification, tibial tubercle-trochlear groove (TT-TG) distance, and patellar height (with Insall-Salvati index) were measured. Pre- and postoperative patellar tilt were compared. Information on outcome scores, ability to return to sports, postoperative recurrent dislocations, and complications was recorded. RESULTS: A total of 42 MPFC-Rs in 39 patients were included. Mean age at surgery was 22.2 ± 7.6 years; 69.2% of patients were female. Mean follow-up was 47.3 ± 20.2 months. Seventy-four percent of cases had Dejour B (19.0%), C (33.3%), and D (21.4%) trochlear dysplasia; mean TT-TG distance was 19.6 ± 3.5 mm, and mean Insall-Salvati index was 1.21 ± 0.17. Mean patellar tilt decreased from 27.6 ± 11.6° to 9.4 ± 6.5° (P < .001). All patients had statistically significant (P < .001) improvement in mean International Knee Documentation Committee (IKDC) (44.9 ± 18.2 to 87.5 ± 6.9), Lysholm (61.4 ± 16.6 to 94.1 ± 6.4), Kujala (56.0 ± 16.8 to 92.9 ± 5.3), and Tegner score (2.7 ± 1.3 to 4.6 ± 1.4). The majority of patients (96.9%) returned to sports, with 90.3% returning to the same or greater level of activity. No postoperative dislocations or subluxations were reported. CONCLUSIONS: Arthroscopically assisted double-bundle MPFC-R is a promising procedure to treat recurrent patellar instability at 2- to 7-year mid-term follow-up, despite the presence of trochlear dysplasia, elevated TT-TG distance and patellar alta. The improvement of IKDC score exceeded the minimal clinically important difference in 95.2% patients, and 66.7% surpassed the patient acceptable symptomatic state based on postoperative IKDC score with no redislocations being reported at latest follow-up. LEVEL OF EVIDENCE: Level IV, case series, retrospective.

Comparison and analysis of skiing injuries at ski resorts in Chongli, China and Japan.

PURPOSE: The rapid development of winter sports requires investigation on injuries in Chongli district, Zhangjiakou city, one of the ski sites of the 2022 Winter Olympics. Careful evaluation is required to observe which injuries are caused under what circumstances, and then we can make corresponding preventive measures and recommendations based on the results. METHODS: In this retrospective study, the data of injury cases at ski resorts in China (Chongli district) and Japan were analyzed to provide a reference for the ongoing injury prevention at ski resorts. We collected data on injuries at Wanlong and Fulong ski resorts in Chongli district during the 2017-2018 and 2018-2019 ski seasons. We referred to the skiing injury report issued in February 2020 of a nationwide ski safety statistical service - Japan Ski Safety Association. The causes of injury and specific injured body parts were analyzed based on the data of Chinese and Japanese ski resorts. Statistical significance has been calculated using the Chi-square test. RESULTS: During the 2019-2020 ski season in Japanese ski resorts, the number of reported injuries per 10,000 skiers was 0.93, of which 457 (17.3%) were over 50 years old, accounting for a large proportion of injuries, meanwhile in Chongli ski resort, the injury rate of skiers aged 50 and over was 7.1%. The knee joint (23.7% at Wanlong ski resort and 28.4% in Japanese ski resorts) was the most injured body part among Chongli and Japanese skiers. Among snowboarders, shoulder joint injury (17.7% in Japanese ski resorts) was the most common, and injury on hands and fingers (16.3% in Wanlong ski resorts) was the most common. Head injury rates are similar in Chongli, China and Japanese ski resorts (8.2% and 8.7%, respectively). CONCLUSION: Our analysis demonstrated that injury data recorded among young skiers was higher in Chinese ski resorts (Chongli district) than that in Japanese ski resorts, and elderly skiers made up a larger proportion of skiing injuries in Japanese resorts. Thus, according to our research, the protection of knee joints, shoulder joints, and hands and fingers should be taken seriously. It should pay attention to the teaching of ski poles (for finger protection), and use protective devices such as knee pads, helmets, etc.

The selection and identification of compound housekeeping genes for quantitative real-time polymerase chain reaction analysis in rat fetal kidney.

During quantitative real-time polymerase chain reaction (RT-qPCR) data analysis, the selection of optimal housekeeping gene is necessary to ensure the accuracy of results. It is noteworthy that housekeeping genes commonly used in adult studies may not be applicable for fetus. However, the stability analysis of housekeeping gene in fetal kidney has not been reported. This study intends to screen the applicable compound housekeeping genes in rat fetal kidney. In this study, eight housekeeping genes used in kidney studies based on literature reports (GAPDH, ACTB, 18S, HPRT, YWHAZ, HMBS, PPIA, and TBP) were selected as the research object. Their expression levels in the rat fetal kidney in physiological condition and the intrauterine growth retardation (IUGR) model induced by prenatal dexamethasone exposure (PDE) (0.2 mg/kg·day from gestation Days 9 to 20) was measured. Furthermore, these eight housekeeping genes were used to conduct relative quantitative analysis of nephrin expression in the fetal kidney in PDE-induced IUGR model, to compare the influence of choosing different housekeeping gene on data analysis of nephrin expression and to verify the reliability of selected compound housekeeping genes. In this study, stable housekeeping genes of fetal kidney tissues in PDE-induced IUGR model were identified: ACTB, GAPDH, TBP, and HMBS for males; ACTB, YWHAZ, and GAPDH for females. Besides, our results suggest that ACTB + GAPDH were the best compound housekeeping genes for normalization analysis in male fetal kidney studies, and ACTB + YWHAZ in females. This study will provide an experimental evidence basis for the selection of housekeeping genes in the RT-qPCR experiment in renal development toxicology-related models.

Arthroscopic modified Broström procedure achieves faster return to sports than open procedure for chronic ankle instability.

PURPOSE: To compare the clinical outcomes, rate of return to sports, postural control, and muscle strength between the arthroscopic and open modified Broström procedure for chronic lateral ankle instability (CLAI) patients. METHODS: From September 2018 to April 2019, 70 patients diagnosed with CLAI were prospectively included with arthroscopic modified Broström procedure (n = 36) and open modified Broström procedure (n = 34). They were evaluated at five time points (preoperation and 3 months, 6 months, 1 year and 2 years postoperatively). The main results examined the rate of return to sports, American Orthopaedic Foot and Ankle Society Score (AOFAS), Foot and Ankle Ability Measure (FAAM), visual analogue scale (VAS), centre of pressure (COP) excursion velocity, time to boundary (TTB), plantar pressure, isokinetic muscle strength and complications. RESULTS: Compared with the open group, the arthroscopic group demonstrated a significantly shorter period of return to the preinjury sport (13.2 ± 2.4 weeks vs. 18.7 ± 3.1 weeks, P = 0.023) and a higher early sport ratio (80.6 vs. 61.8%, P = 0.011) combined with better FAAM sports and AOFAS at 3 months and 6 months postoperatively and VAS at 3 months postoperatively. In addition, better anterior-posterior postural control stability, less time to peak force under lateral hindfoot and better dorsiflexion strength were shown in the arthroscopic group at 6 months postoperatively. No significant difference was found in clinical scores, posture control or muscle strength at the 1- or 2-year follow-up between the two groups. CONCLUSIONS: Shorter period and higher rates of return to sport activities and better clinical scores, posture control and muscle strength were achieved in the arthroscopic group at 6 months postoperatively, and no clinical differences were found between arthroscopic and open modified Broström procedure 1 year or 2 years postoperatively. Arthroscopic modified Broström procedure is a reliable procedure for CLAI injuries with the demand for fast exercise recovery. CLINICAL REGISTRATION: ChiCTR1900023999. LEVEL OF EVIDENCE: II.

Meniscus repair with simultaneous anterior cruciate ligament reconstruction: Clinical outcomes, failure rates and subsequent processing.

PURPOSE: To retrospectively analyze the clinical outcomes of meniscus repair with simultaneous anterior cruciate ligament (ACL) reconstruction and explore the causes of failure of meniscus repair. METHODS: From May 2013 to July 2018, the clinical data of 165 patients who were treated with meniscus surgery and simultaneous ACL reconstruction, including 69 cases of meniscus repair (repair group) and 96 cases of partial meniscectomy (partial meniscectomy group) were retrospectively analyzed. The exclusion criteria were as follows: (1) ACL rupture associated with fracture, collateral ligament injury, or complex ligament injury; (2) a history of knee surgery; or (3) a significant degree of osteoarthritis. The 69 patients in the repair group were divided into the non-failure group (62 cases) and the failure group (7 cases) depending on the repair effect. Postoperative outcomes of the repair group and the partial meniscectomy group were compared. General conditions and postoperative outcomes of the failure group and the non-failure group were compared. During the median follow-up period of 28 months (range, 4 - 65 months) after the second arthroscopy, postoperative outcomes of seven patients in the failure group were summarized. SPSS 25.0 statistical software was used for statistical analysis. A p value less than 0.05 was considered statistically significant. RESULTS: Seven patients in the failure group who underwent the second arthroscopy were followed up for (30 ± 17.4) months and their postoperative outcomes were summarized. Compared with the partial meniscectomy group, the International Knee Documentation Committee scores of patients in the repair group improved significantly (p = 0.031). Compared with the non-failure group, more patients in the failure group were younger than 24 years (p = 0.030). The median follow-up period was 39.5 months. All patients recovered well after subsequent partial meniscectomy and relieved clinical symptoms. Visual analog scale scores decreased significantly (p = 0.026), and the International Knee Documentation Committee and Lysholm scores improved significantly (p = 0.046 for both). CONCLUSION: The failure rate of meniscus repair in this study was 10.1% (7/69), all of which were medial meniscus tears. However, the surgical outcomes of ACL reconstruction were not affected, and there might be a role for graft protection. Therefore, meniscus retears can be successful treated by performing subsequent partial meniscectomy in patients with repair failure.

Secondary Effects of the Rupture and Reconstruction of the Interosseous Talocalcaneal Ligament on the Peritalar Joints.

BACKGROUND The interosseous talocalcaneal ligament (ITCL) is the main soft-tissue contributor to subtalar joint stability. The role of ITCL reconstruction in retaining this stability is minimally reported. Therefore, we conducted this study to investigate the effects of rupture and reconstruction of the ITCL on the subtalar and peritalar joints. MATERIAL AND METHODS This experimental study randomly divided 72 rabbits into 3 equal groups of 24 rabbits each. Group I underwent reconstruction surgery, group II underwent resection, and group III was the control group. The cartilages between the talocrural and calcaneocrural joints, and between the subtalar and talonavicular joints on both sides were assessed by gross observation, ink staining, histology, and immunohistochemistry at weeks 4, 8, 16, and 32, postoperatively. RESULTS In group II, the quantitative ink staining analysis revealed degeneration of the articular cartilages on the talonavicular joint (T=2.070, P=0.038) and the posterior subtalar joint (T=2.121, P=0.034) compared with the 2 sides of the same rabbit at 4 and 8 postoperative weeks. Comparing the operated sides of all the groups showed the posterior subtalar joints (Hc=9.563, P=0.008) and talonavicular joints (Hc=9.714, P=0.008) had an obvious difference at postoperative week 4; and in the calcaneocrural joints (Hc=6.750, P=0.034), it was noticed at postoperative week 8. Histology and immunohistochemistry findings confirm these observations. CONCLUSIONS An ITCL resection can lead to the progressive degeneration of the talonavicular and posterior subtalar joints, while an ITCL reconstruction can be beneficial in restoring the stability of these joints, preventing or postponing their degeneration, and protecting the articular cartilages.

Characteristics and predictors of muscle strength deficit in mechanical ankle instability.

PURPOSE: Muscle strength training is a common strategy for treating chronic ankle instability (CAI), but the effectiveness decreases for mechanical ankle instability (MAI) patients with initial severe ligament injuries. The purpose of this study was to investigate the characteristics and the potential predictors of muscle strength deficit in MAI patients, with a view to proposing a more targeted muscle strength training strategy. METHODS: A total of 220 MAI patients with confirmed initial lateral ankle ligament rupture and a postinjury duration of more than 6 months were included. All patients underwent a Biodex isokinetic examination of the ankle joints of both the affected and unaffected sides. Then, the associations between the limb symmetry index (LSI) (mean peak torque of the injury side divided by that of the healthy side) and the patients' sex, body mass index, postinjury duration, presence of intra-articular osteochondral lesions, presence of osteophytes and ligament injury pattern (i.e., isolated anterior talofibular ligament (ATFL) injury or combined with calcaneofibular ligament injury) were analysed. RESULTS: There was significantly weaker muscle strength on the affected side than on the unaffected side in all directions (p < 0.05). The LSI in plantar flexion was significantly lower than that in dorsiflexion at 60°/s (0.87 vs 0.98, p < 0.001). A lower LSI in eversion was significantly correlated with female sex (0.82 vs 0.94, p = 0.016) and isolated ATFL injury (0.86 vs 0.95, p = 0.012). No other factors were found to be associated with muscle strength deficits. CONCLUSION: MAI patients showed significant muscle strength deficits on the affected side, especially in plantar flexion. There were greater strength deficits in eversion in females and individuals with an isolated ATFL injury. Thus, a muscle strength training programme for MAI patients was proposed that focused more on plantar flexion training and eversion training for females and those with an isolated ATFL injury.

[The role of miRNA in kidney development].

MicroRNAs (miRNAs) are endogenous small non-coding RNAs (19-25 nucleotides) that negatively regulate gene expression at the post transcriptional level by binding to complementary target sequences in the target mRNA. miRNAs play an important role in a wide range of biological processes, including organ development. Recent studies have shown that some miRNAs are highly expressed in the kidney and are closely related to kidney development and diseases, suggesting that miRNAs are important regulators in kidney physiology and pathology. This review will focus on the research progress of miRNA in kidney development, and discuss the role of miRNAs in the occurrence and development of renal dysplasia, which will provide a reference for the diagnosis and research of diseases related to kidney development.

The effect of cyclic knee motion on the elongation of four-strand hamstring autograft in anterior cruciate ligament reconstruction: an in-situ pilot study.

BACKGROUND: Pretension of the viscoelastic graft by cyclic knee motion has been confirmed to decrease the graft creep and improve the outcome of anterior cruciate ligament (ACL) reconstruction. The purpose of the present study was to investigate the effect of cyclic knee motion on the elongation of the four-strand hamstring tendon autograft in situ and to explore the stable level cycle, in which the tendon length achieved a stable level. METHODS: The study was performed with 53 consecutive patients undergoing transtibial ACL reconstruction with four strand hamstring tendon from Aug 2013 to Apr 2015. 43 males and 10 females were included with mean age of 29 ± 10 years. The pretension of the tendons was operated by cyclical knee motion ranging from 0 to 110°after the femoral fixation with Endo-button. The tendon length after 10, 20, 30 and 40 cycles was measured respectively and compared by repeated measure ANOVA. Then multivariate logistic regression was used to investigate the effect of the patients' parameters (age, gender, height, body weight, tendon length, etc.) on the elongation of the graft and the stable level cycle. RESULTS: The mean lengthening of the graft at 10, 20, 30 and 40 times was 3.0 ± 1.4 mm, 4.3 ± 1.5 mm, 4.8 ± 1.7 mm and 4.8 ± 1.8 mm respectively. No significant correlation was found between the elongation and the patients' parameters. There was significant difference of the tendon length from 0 to 30 cycles (F = 264.8, df = 1.95, p<0.001). However, the tendon length achieved a stable level after 30 cycles and the median elongation from 30 cycles to 40 cycles was 0 (0-1) mm with no significant difference (F = 2.039, p = 0.159). The male and female tendon length achieved to a stable level at 20 cycles and 30 cycles respectively but with no significant difference (p = 0.074). CONCLUSIONS: The four-strand hamstring tendon was elongated after cyclic knee motion and the elongation achieved a stable level after 30 cycles for the transtibial technique. Both of the tendon elongation and the stable level cycle were not correlated with patients' gender, age, preoperative duration, graft diameter and length.

Autophagy as a compensation mechanism participates in ethanol-induced fetal adrenal dysfunction in female rats.

Autophagy plays a vital role in embryonic development and cell differentiation. Our previous study demonstrated that prenatal ethanol exposure (PEE) resulted in intrauterine growth retardation (IUGR) and adrenal developmental toxicities in rat offspring. The present study focused on PEE-induced autophagy as an underlying mechanism and its biological significance in female fetal rats. Female fetuses in the PEE group exhibited lower body weights and suffered adrenal structural abnormalities compared to the controls. Cell proliferation was inhibited, the insulin-like growth factor 1 (IGF1) pathway was reduced, and autophagy was activated in the glands of female fetal rats. Ethanol increased the ratio of microtubule-associated protein light chain 3 beta-II (LC3ß-II) to LC3ß-I in vitro, and it reduced cortisol levels in time- and concentration-dependent manners in human adrenocortical carcinoma cells (NCI-H295A). Bafilomycin A1 inhibited autophagy, steroidogenic factor 1 (SF1) protein and steroidogenesis in the present study. Rapamycin with ethanol up-regulated autophagy and SF1 expression and activated steroidogenesis when compared with ethanol alone. In addition, ethanol inhibited IGF1 receptor (IGF1R) and phospho-mTOR (Ser2448) expression in a concentration-dependent manner. These results demonstrate that PEE activated autophagy in fetal adrenal glands, and the underlying mechanism may be associated with inhibition of the IGF1R/phospho-mTOR (Ser2448) pathway. Autophagy may be a compensatory mechanism for the PEE-induced inhibition of fetal adrenal steroidogenesis to maintain fetal adrenal development.

Developmental disorder of podocytes and the related renal diseases.

Podocyte is one of the main components of glomerular filtration barrier in the kidney; the loss or dysfunction of podocyte could impair the functions of glomerular filtration barrier, leading to development of various renal diseases. Podocyte is a terminally differentiated cell, and thus does not possess any proliferative properties. Accordingly, its number and contribution to renal function are initially determined by its normal development. Information from the literature and results of our research indicate that genetic factors or prenatal adverse environment could cause developmental retardation of podocytes, thereby suggesting the potential fetal developmental origin(s) of kidney diseases, and involvement of epigenetic mechanisms in the regulation of key genes in podocyte development. In this review, we provide a brief overview on the podocyte normal development; discussion on the potential pathogenic mechanisms for developmental disorders of podocytes; as well as renal diseases associated with podocyte developmental retardation. We aim to provide some insights in articulating the strategies for diagnosis and treatments of renal diseases associated with podocyte developmental abnormalities.

Indole-3-Carbinol Induces Apoptosis of Hepatic Stellate Cells through K63 De-Ubiquitination of RIP1 in Rats.

BACKGROUND/AIMS: The apoptosis of activated hepatic stellate cells (HSCs) is the central event in the reversal of liver fibrosis. K63 de-ubiquitinated receptor-interacting protein (RIP)1 promotes apoptosis in tumor necrosis factor (TNF)-α signaling pathway. In the previous study, we have proved that indole-3-carbinol (I3C) could reverse different models of liver fibrosis in rats, but the mechanism is still unclear. Thus, the present research aimed to demonstrate the induction of I3C on apoptosis of HSCs and the underlying molecular mechanism. METHODS: HSC-T6, an immortalized rat liver stellate cell line, was treated for 24 hours with 25, 50 and 100 µM of I3C. The apoptosis of HSC-T6 was analyzed by flow cytometric analysis, acridine orange staining and RT-PCR, respectively. K63 de-ubiquitination of RIP1 and the expression of ubiquitin ligases and deubiquitinases were analyzed by Co-IP assay and western blot. Knockdown of deubiquitinases was undertaken by small interfering RNA (siRNA). RESULTS: The results of flow cytometric analysis indicated that the apoptotic rate of HSC-T6 was induced by I3C in a dose-dependent manner. Observation by acridine orange staining exhibited nuclear condensation and apoptotic bodies in I3C treated cells. Consistently, the expression ratio of Bax/bcl-2 was markedly increased by I3C. These results indicated that I3C could significantly induce apoptosis of HSC-T6 cells. Furthermore, Co-IP assay revealed K63 de-ubiquitination of RIP1 by I3C, associated with the induction of caspase 8. Although I3C had no effect on the expression of ubiquitin ligases cellular inhibitor of apoptosis 1/2 (cIAP1/2), the protein level of deubiquitinase cylindromatosis (CYLD) was significantly induced by I3C. Moreover, CYLD silencing reversed the pro-apoptosis induction effect of I3C and reduced the expression ratio of Bax/bcl-2 in HSC-T6 cells. CONCLUSION: These results demonstrated that I3C could induce apoptosis of HSC through RIP1 K63 de-ubiquitination by upregulating deubiquitinase CYLD.

OsCERK1 and OsRLCK176 play important roles in peptidoglycan and chitin signaling in rice innate immunity.

Microbe-associated molecular pattern (MAMP)-triggered immunity plays critical roles in the basal resistance defense response in plants. Chitin and peptidoglycan (PGN) are major molecular patterns for fungi and bacteria, respectively. Two rice (Oryza sativa) lysin motif-containing proteins, OsLYP4 and OsLYP6, function as receptors that sense bacterial PGN and fungal chitin. These membrane receptors, which lack intracellular kinase domains, likely contain another component for transmembrane immune signal transduction. Here, we demonstrate that the rice LysM receptor-like kinase OsCERK1, a key component of the chitin elicitor signaling pathway, also plays an important role in PGN-triggered immunity in rice. Silencing of OsCERK1 suppressed PGN-induced (and chitin-induced) immunity responses, including reactive oxygen species generation, defense gene expression, and callose deposition, indicating that OsCERK1 is essential for both PGN and chitin signaling initiated by OsLYP4 and OsLYP6. OsLYP4 associated with OsLYP6 and the rice chitin receptor chitin oligosaccharide elicitor-binding protein (CEBiP) in the absence of PGN or chitin, and treatment with PGN or chitin led to their disassociation in vivo. OsCERK1 associated with OsLYP4 or OsLYP6 when induced by PGN but it associated with OsLYP4, OsLYP6, or CEBiP under chitin treatment, suggesting the presence of different patterns of ligand-induced heterooligomeric receptor complexes. Furthermore, the receptor-like cytoplasmic kinase OsRLCK176 functions downstream of OsCERK1 in the PGN and chitin signaling pathways, suggesting that these MAMPs share overlapping intracellular signaling components. Therefore, OsCERK1 plays dual roles in PGN and chitin signaling in rice innate immunity and as an adaptor involved in signal transduction at the plasma membrane in conjunction with OsLYP4 and OsLYP6.

Lysin motif-containing proteins LYP4 and LYP6 play dual roles in peptidoglycan and chitin perception in rice innate immunity.

Plant innate immunity relies on successful detection of microbe-associated molecular patterns (MAMPs) of invading microbes via pattern recognition receptors (PRRs) at the plant cell surface. Here, we report two homologous rice (Oryza sativa) lysin motif-containing proteins, LYP4 and LYP6, as dual functional PRRs sensing bacterial peptidoglycan (PGN) and fungal chitin. Live cell imaging and microsomal fractionation consistently revealed the plasma membrane localization of these proteins in rice cells. Transcription of these two genes could be induced rapidly upon exposure to bacterial pathogens or diverse MAMPs. Both proteins selectively bound PGN and chitin but not lipopolysaccharide (LPS) in vitro. Accordingly, silencing of either LYP specifically impaired PGN- or chitin- but not LPS-induced defense responses in rice, including reactive oxygen species generation, defense gene activation, and callose deposition, leading to compromised resistance against bacterial pathogen Xanthomonas oryzae and fungal pathogen Magnaporthe oryzae. Interestingly, pretreatment with excess PGN dramatically attenuated the alkalinization response of rice cells to chitin but not to flagellin; vice versa, pretreatment with chitin attenuated the response to PGN, suggesting that PGN and chitin engage overlapping perception components in rice. Collectively, our data support the notion that LYP4 and LYP6 are promiscuous PRRs for PGN and chitin in rice innate immunity.

Low functional programming of renal AT2R mediates the developmental origin of glomerulosclerosis in adult offspring induced by prenatal caffeine exposure.

Our previous study has indicated that prenatal caffeine exposure (PCE) could induce intrauterine growth retardation (IUGR) of offspring. Recent research suggested that IUGR is a risk factor for glomerulosclerosis. However, whether PCE could induce glomerulosclerosis and its underlying mechanisms remain unknown. This study aimed to demonstrate the induction to glomerulosclerosis in adult offspring by PCE and its intrauterine programming mechanisms. A rat model of IUGR was established by PCE, male fetuses and adult offspring at the age of postnatal week 24 were euthanized. The results revealed that the adult offspring kidneys in the PCE group exhibited glomerulosclerosis as well as interstitial fibrosis, accompanied by elevated levels of serum creatinine and urine protein. Renal angiotensin II receptor type 2 (AT2R) gene expression in adult offspring was reduced by PCE, whereas the renal angiotensin II receptor type 1a (AT1aR)/AT2R expression ratio was increased. The fetal kidneys in the PCE group displayed an enlarged Bowman's space and a shrunken glomerular tuft, accompanied by a reduced cortex width and an increase in the nephrogenic zone/cortical zone ratio. Observation by electronic microscope revealed structural damage of podocytes; the reduced expression level of podocyte marker genes, nephrin and podocin, was also detected by q-PCR. Moreover, AT2R gene and protein expressions in fetal kidneys were inhibited by PCE, associated with the repression of the gene expression of glial-cell-line-derived neurotrophic factor (GDNF)/tyrosine kinase receptor (c-Ret) signaling pathway. These results demonstrated that PCE could induce dysplasia of fetal kidneys as well as glomerulosclerosis of adult offspring, and the low functional programming of renal AT2R might mediate the developmental origin of adult glomerulosclerosis.

[Effect of anterior cruciate ligament rupture on hamstring: quadriceps ratio during isokinetic knee extension and flexion at 30 degrees of flexion].

OBJECTIVE: To evaluate the change in hamstring (H):quadriceps (Q) ratio following anterior cruciate ligament (ACL) rupture during isokinetic knee extension and flexion at 30 degrees of flexion which is important for knee dynamic function. METHODS: A study was performed in 25 male complete unilateral ACL ruptures. Isokinetic concentric and eccentric quadriceps and hamstring muscle tests in both the deficient knees and intact knees were performed at 60°/s, respectively. At 30 degrees of flexion, the average torque of quadriceps and hamstring, Qe:Qc ratios (ratios of eccentric quadriceps to concentric quadriceps muscle torque), He:Hc ratios (eccentric hamstring to concentric hamstring), Hc:Qc ratios (concentric hamstring to concentric quadriceps), He:Qc ratios (eccentric hamstring to concentric quadriceps), and Hc:Qe ratios (concentric hamstring to eccentric quadriceps) were calculated. Wilcoxon matched-pairs signed-ranks test was used. RESULTS: At 30 degrees of knee flexion, a significant reduction (P<0.05) in the average torque of quadriceps was observed at concentric and eccentric 60°/s produced by the deficient-side compared with the intact side. In addition, Hc:Qc, He:Qc, and Qe:Qc significantly increased on the ACL-deficient side. CONCLUSION: The change in H :Q ratio in the mode of isokinetic 60°/s at 30 degrees of knee flexion might therefore be a new tool to objectively document muscle function in ACL-deficient knee.

Prospective randomized comparison of anatomic single- and double-bundle anterior cruciate ligament reconstruction.

PURPOSE: To determine if anatomic double-bundle anterior cruciate ligament (ACL) reconstruction is superior to anatomic single-bundle reconstruction in restoring the stabilities and functions of the knee joint. METHODS: A prospective randomized clinical study was done to compare the results of 32 cases of anatomic single-bundle ACL reconstruction and 34 cases of anatomic double-bundle ACL reconstruction with average follow-up of 16.3 ± 3.1 months. Tunnel placements of all the cases were measured on 3D CT. Clinical results were collected after reconstruction; graft's appearance, meniscus status and cartilage state under arthroscopy were compared and analysed too. RESULTS: Tunnel placements, confirmed with 3D CT, were in the anatomic positions as described in literature both in SB and DB group. No differences were found between SB and DB groups in clinical outcome scores, pivot shift test and KT 1000 measurements (average side-to-side difference for anterior tibial translation was 0.7 mm in SB group and 1.0 mm in DB group). More than 70 % of the single-bundle graft and AM bundle graft in DB group appeared excellent, but only 44.1 % of PL bundle grafts in DB group were excellent and 11.8 % were in poor state. No new menisci tear was found either in SB or DB group, however, in DB group cartilage damages in medial patella-femoral joint occurred in 38.2 % cases. This rate was significantly higher than in the SB group which is only 9.3 %. CONCLUSION: Both single- and double-bundle anatomic ACL reconstruction can restore the knee's stability and functions very well. However, more incidences of poor PL status and medial patellar-femoral cartilage damage may occur in double-bundle ACL reconstruction.