Neuroprotective effect of triptolide on neuronal inflammation in rats with mild brain injury.

Concussions sustained while playing sports are a prominent cause of mild traumatic brain injury (mTBI), which is prevalent among teenagers. The early and intermediate stages of mild traumatic brain injury (mTBI) can be characterized by inflammation, neurodegeneration, and brain tissue edema, which can lead to permanent brain damage. The present study investigated the therapeutic effects of triptolide in mTBI and brain damage recovery. After building mTBI model in male rat, triptolide administrated daily for 1 week in the treated group. On day 3 and day 7 of administration, hippocampus tissues were collected to evaluate inflammation and autophagy in the brain. The expressions of inflammatory factors interleukin (IL)-1ß and tumor necrosis factor-alpha in serum were downregulated, while IL-10 expression was upregulated when compared with the mTBI group on day 3 and day 7. The expression of IL-10 on day 7 was higher than on day 3. Quantitative polymerase chain reaction (qPCR) analysis of inflammatory-related factors (i.e., Il-1ß and nuclear factor-κB (Nf-κb), and western blot as well as immunofluorescence staining of autophagy-related proteins (i.e., LC3B) and aquaporin (AQP 4) showed lower expression on day 3 and day 7 in the triptolide-treated group. Moreover, NeuN immunostaining, and hematoxylin and eosin (HE) staining for hippocampus region revealed that the triptolide-treated group showed a decrease in damaged cells. Our findings emphasize the effectiveness of triptolide therapy after mild traumatic brain injury via modulating autophagy, attenuating inflammation and reduces edema by decreasing AQP 4 expression.

"Adjacent Bed Effect" of Total Knee Arthroplasty Patients During the Perioperative Period.

BACKGROUND: Knee osteoarthropathy is one of the most common degenerative joint diseases in the elderly, total knee arthroplasty (TKA) is the most commonly used treatment for end-stage knee osteoarthropathy. Negative emotions such as anxiety have been extensively documented in knee osteoarthropathy patients. AIM: This study aimed to investigate the Emotional Contagion during hospitalization in patients undergoing TKA. METHODS: Eligible subjects were divided into three case groups according to their anxiety states and bed arrangement. All subjects underwent a unilateral, cemented TKA under general anesthesia. Post-operative recovery outcomes including pain, pain behavior and physical function were recorded pre-operation, 1-day, 1 week, 2-weeks, 1-month and 3-months post-operation. RESULTS: A total of 38 subjects were included in the final analysis. Subjects with anxiety had higher Visual Analogue Scale pain scores, PROMIS-Pain Behavior scores than subjects without anxiety in the Contagion Group preoperation (p ≤ .05). Non-anxiety subjects hospitalized in beds physically adjacent to anxiety subjects experienced more severe pain and poorer function (p ≤ .05). After discharge, all clinical outcomes gradually became lower than anxiety subjects in the Contagion Group, reaching levels similar to non-anxiety subjects in the No Contagion Group within 1 month (p>.05). CONCLUSIONS: This study showed that patients with anxiety may have an "Adjacent Bed Effect" on patients with TKA in the adjacent bed, which may be associated with poorer postoperative recovery, including pain and physical function. We speculate this phenomenon can be effectively avoided by the nursing team through accurately assessing psychological status and reasonable bed arrangements in the inpatient assessment phase.

[Application of femoral I.D.E.A.L localization in anterior cruciate ligament reconstruction with single bundle anterior cruciate ligament].

OBJECTIVE: To evaluate the effect of femoral I.D.E.A.L localization in single bundle anterior cruciate ligament reconstruction (ACLR). METHODS: From January 2019 to October 2022, 122 patients with anterior cruciate ligament injury were treated with ACLR, including 83 males and 39 females. The age ranged from 23 to 43 years old, with an average of (32.19 ±8.55) years old. The course of disease ranged from 1 week to 6 months. According to the different surgical schemes, the patients were divided into two groups, namely the traditional group, which adopted the over-the-top femoral lateral positioning scheme, including 64 patients. The I.D.E.A.L group adopted the I.D.E.A.L femoral lateral positioning scheme, including 58 patients. The patient has pain and dysfunction of knee joint before operation. MRI of knee joint indicates anterior cruciate ligament injury. The visual analogue scale(VAS), International Knee Documentation Committee(IKDC) scoring system and Lysholm scoring system were used to evaluate the knee joint function of the patient. KT-2000 was used to detect the recovery of knee joint after operation and to count the postoperative complications. RESULTS: The wounds healed well after operation. One hundred and twenty-tow patients were followed up for 15 to 46 months, with an average of (25.45±9.22) months. The knee joint stability of patients after operation was significantly increased. The VAS at 1 day and 1 week after operation of patients in the I.D.E.A.L group was significantly lower than that in the traditional group(P<0.05). The IKDC score and Lysholm score of patients in the I.D.E.A.L group were significantly higher than those in the traditional group(P<0.05). In the traditional group, there were 6 cases of short-term (<1 month) complications and 19 cases of long-term (≥1 month)complicatios. In the I.D.E.A.L group, there were 3 cases of short-term complications and 7cases of long-term complications(P<0.05). CONCLUSION: The single bundle anterior cruciate ligament reconstruction and femoral I.D.E.A.L positioning can achieve better early postoperative effect and reduce early postoperative pain.

Barefoot walking is more stable in the gait of balance recovery in older adults.

BACKGROUND: Perturbation-based balance training on a treadmill is an emerging method of gait stability training with a characteristic task nature that has had positive and sustained effects on balance recovery strategies and fall reduction. Little is known about the effects produced by shod and barefoot walking. We aimed to investigate which is more appropriate, shod or barefoot walking, for perturbation-based balance training in older adults. METHODS: Fourteen healthy older adults (age: 68.29 ± 3.41 years; body height: 1.76 ± 0.10 m; body mass: 81.14 ± 14.52 kg) performed normal and trip-like perturbed walking trials, shod and barefoot, on a treadmill of the Gait Real-time Analysis Interactive Lab. The marker trajectories data were processed by Human Body Model software embedded in the Gait Offline Analysis Tool. The outcomes of stride length variability, stride time variability, step width variability, and swing time variability were computed and statistically analyzed by a two-way repeated-measures analysis of variance (ANOVA) based on gait pattern (normal gait versus perturbed recovery gait) and footwear condition (shod versus barefoot). RESULTS: Footwear condition effect (p = 0.0310) and gait pattern by footwear condition interaction effect (p = 0.0055) were only observed in swing time variability. Gait pattern effects were detected in all four outcomes of gait variability. CONCLUSIONS: Swing time variability, independent of gait speed, could be a valid indicator to differentiate between footwear conditions. The lower swing time variability in perturbed recovery gait suggests that barefoot walking may be superior to shod walking for perturbation-based balance training in older adults.

Compensatory Responses During Slip-Induced Perturbation in Patients With Knee Osteoarthritis Compared With Healthy Older Adults: An Increased Risk of Falls?

Background: Functional impairment of the knee joint affected by osteoarthritis and loss of muscle strength leads to a significant increase in the number of falls. Nevertheless, little is known about strategies for coping with gait perturbations in patients with knee osteoarthritis (KOA). Thus, this study aimed to examine the compensatory strategies of patients with KOA in response to a backward slip perturbation compared with healthy older adults. Methods: An automated perturbation program was developed by using D-Flow software based on the Gait Real-time Analysis Interactive Lab, and an induced backward slip perturbation was implemented on nine patients with severe KOA (68.89 ± 3.59 years) and 15 age-matched healthy older adults (68.33 ± 3.29 years). Step length, gait speed, range of motion, vertical ground reaction forces, lower extremity joint angles, and joint moments were computed and analyzed. Results: Compared with older adults, patients with KOA had significantly lower step length, gait speed, and vertical ground reaction forces in both normal walking and the first recovery step following backward slip perturbations. Inadequate flexion and extension of joint angles and insufficient generation of joint moments predispose patients with KOA to fall. Hip extension angle and flexion moment, knee range of motion, and vertical ground reaction forces are key monitoring variables. Conclusion: The risk of falls for patients with KOA in response to backward slip perturbations is higher. Patients with KOA should focus not only on quadriceps muscle strength related to knee range of motion but also on improving hip extensor strength and activation through specific exercises. Targeted resistance training and perturbation-based gait training could be better options.

[Repair of osteoarthritis in animal model with exosomes derived from BMSCs transfected by the siRNA -Piezo1 through CT navigation].

OBJECTIVE: To investigate the effect of the exosomes from bone marrow mesenchymal stem cells (BMSCs) transfected with silence plasmid of Piezol small interference RNA (siRNA)on osteoarthritis (OA) animal model. METHODS: Twenty male SD rats with specific pathogen free (SPF) were selected, ranging in age from 5.46 to 6.96 months, with a mean of (6.21± 0.75) months;and ranging in weight from 385.76 g to 428.66 g, with a mean of (407.21±21.45) g. BMSCs were extracted. The siRNA silencing plasmid of piezo1 was constructed by siRNA technology. After lentivirus was transfected into BMSCs, the exosomes were extracted. At the cellular level, BMSCs were divided into blank plasmid group and siRNA silencing plasmid group according to whether siRNA-Piezo1 was transfected or not. The osteogenic induction ability of siRNA-Piezo1 on BMSCs was detected by RT-PCR and Western blot. At the animal model level, the OA model was established by surgical resection of cruciate ligament of knee joint.According to different treatment schemes, SD rats were divided into 4 groups:blank control group, model group, BMSCs group and exosome group. SD rats in the blank control group were not treated. In the model group, the cruciate ligaments of rats were excised and OA animal model was established. In BMSCs group, BMSCs were injected into knee joint under CT guidance after OA model establishment, and the cell volume was 5×105/ml, loading amount of 2 ml, twice a week for 4 weeks. In the exosome group, 100 µg exosomes from siRNA BMSCs were added twice a week for 4 weeks after OA model establishment. The cartilage of the animal model was detected by hematoxylin eosin (HE) staining and safranin solid green staining, and quantified by the modified Minkin score and the score of the international society for osteoarthritis research (OARSI). Reverse transcription polymerase chain reaction (RT-PCR) was used to detect the relative mRNA expression level of aggrecan type II collagen in cartilage. RESULTS: The lentivirus transfection efficiency was(92.11±4.22)%. RT-PCR showed that the relative expression of Piezo1 mRNA in blank plasmid group was 1.07±0.06, which was significantly different from that of 0.31±0.01 in siRNA silencing plasmid group (t=2.907, P<0.05). The results of HE staining and safranine solid green staining showed that there was cartilage structure and smooth cartilage surface in the knee joint of SD rats in the blank control group. The knee joint structure in the model group had been completely destroyed, the knee joint cartilage structure in the BMSCs group was not clear, and there were subchondral bone components in the OA rats in the exosomes group. There was significant difference between the modified Minkin score of HE staining and the OARSI score of safranin solid green staining (F=15.903, 19.005;P<0.05). Among them, the repair effect of exosome group was significantly better than that of BMSCs group and model group (P<0.05). RT-PCR results showed that the relative expression of aggrecan mRNA in BMSCs group was significantly higher than that in model group (P< 0.05), and the relative expression of aggrecan mRNA in exosome group was higher than that in BMSCs group and model group (P<0.05). The relative expression of CollagenⅡmRNA in BMSCs group was higher than that in model group (P<0.05), and the relative expression of CollagenⅡmRNA in exosomes group was higher than that in BMSCs group and model group (P<0.05). CONCLUSION: Piezo1 siRNA silencing vector can promote the differentiation of BMSCs into chondrocytes and effectively inhibit the progression of OA, so as to delay the disease of OA.

Erratum: Dehydrocostus lactone, a natural sesquiterpene lactone, suppresses the biological characteristics of glioma, through inhibition of the NF-κB/COX-2 signaling pathway by targeting IKKß.

[This corrects the article on p. 1270 in vol. 7, PMID: 28670490.].

MiRNA-1271-5p regulates osteogenic differentiation of human bone marrow-derived mesenchymal stem cells by targeting forkhead box O1 (FOXO1).

Forkhead box O1 (FOXO1) is a key regulator of osteogenesis. The aim of this study was to identify the mechanisms of microRNAs (miRNAs) targeting FOXO1 in osteogenic differentiation of human bone marrow mesenchymal stem cells (hMSCs). Three miRNA target prediction programs were used to search for potential miRNAs that target FOXO1. Quantitative real-time polymerase chain reaction was conducted to detect the expression of miR-1271-5p and FOXO1 during osteogenic differentiation. Target gene prediction and screening, luciferase reporter assay was used to verify the downstream target gene of miR-1271-5p. The expression levels of FOXO1 and Runx2 were detected by RT-qPCR and Western blot analysis. Alkaline phosphatase (ALP) activity and matrix mineralization were detected by biochemical methods. The expression levels of Runx2, ALP, and osteocalcin were detected by RT-qPCR. Our results showed that miR-1271-5p was downregulated during osteogenic induction. And the expression levels of miR-1271-5p were higher in osteoporotic tissues than that in adjacent nonosteoporotic tissues. The expression levels of FOXO1 were lower in osteoporotic tissues than that in adjacent nonosteoporotic tissues. And a negative correlation was found between miR-1271-5p and FOXO1 in osteoporotic tissues. Overexpression of miR-1271-5p downregulated FOXO1 and inhibited osteogenic differentiation in hMSCs. Overexpression of miR-1271-5p downregulated the expression of osteogenic markers and reduced ALP activity. In addition, ectopic expression of FOXO1 reversed the effect of miR-1271-5p on osteogenic differentiation. In conclusion, miR-1271-5p functioned as a therapeutic target of osteogenic differentiation in hMSCs by inhibiting FOXO1, which provides valuable insights into the use of miR-1271-5p as a target in the treatment of osteoporosis and other bone metabolic diseases.

Comparison of postoperative effectiveness of less invasive short external rotator sparing approach versus standard posterior approach for total hip arthroplasty.

BACKGROUND: Most of the studies assessing the corrective posterior total hip arthroplasty (THA) mainly focused on the mini-incision approach. Studies exploring the short external rotator sparing approach are rare. Therefore, this study aimed to compare the effectiveness of standard posterior approach and short external rotator sparing approach. METHODS: This prospective observational study included 126 patients who underwent THA in June 2017-June 2018. Patients were assigned to standard (standard posterior approach) and corrective (short external rotator sparing approach) groups based on the surgical method. Surgical data were recorded postoperatively. Postoperative hip joint recovery was assessed using the times to ambulation and independent stair use, and Western Ontario and McMaster Universities Osteoarthritis Index (WOMAC) score, Harris score, and Oxford hip score (OHS) at 2 and 8 postoperative weeks. The visual analog scale (VAS) was used for postoperative pain assessment. RESULTS: Postoperative changes of creatine kinase (CK), myoglobin, CRP, and prosthesis position were similar in both groups (P > 0.05). However, intraoperative blood loss (P < 0.001) and postoperative 6-h drainage volume (P = 0.03), hospital stay, blood transfusion rate, and times to ambulation and independent stair use were significantly reduced in the corrective group. Postoperatively, Oxford, and WOMAC scores significantly decreased in both groups. After surgery, the VAS score was more overtly decreased in the corrective group compared with the standard group. CONCLUSIONS: This study concluded that the less invasive short external rotator sparing approach for THA caused less damage, reducing perioperative blood loss, shortening functional recovery time, maintaining prosthesis stability, and improving postoperative pain.

CRISPR/Cas9 facilitates genomic editing for large-scale functional studies in pluripotent stem cell cultures.

Pluripotent stem cell (PSC) cultures form an integral part of biomedical and medical research due to their capacity to rapidly proliferate and differentiate into hundreds of highly specialized cell types. This makes them a highly useful tool in exploring human physiology and disease. Genomic editing of PSC cultures is an essential method of attaining answers to basic physiological functions, developing in vitro models of human disease, and exploring potential therapeutic strategies and the identification of drug targets. Achieving reliable and efficient genomic editing is an important aspect of using large-scale PSC cultures. The CRISPR/Cas9 genomic editing tool has facilitated highly efficient gene knockout, gene correction, or gene modifications through the design and use of single-guide RNAs which are delivered to the target DNA via Cas9. CRISPR/Cas9 modification of PSCs has furthered the understanding of basic physiology and has been utilized to develop in vitro disease models, to test therapeutic strategies, and to facilitate regenerative or tissue repair approaches. In this review, we discuss the benefits of the CRISPR/Cas9 system in large-scale PSC cultures.

Downregulation of microRNA-23b-3p alleviates IL-1ß-induced injury in chondrogenic CHON-001 cells.

BACKGROUND: Osteoarthritis (OA) is a common joint disease, which is characterized by degradation of articular cartilage. Evidence indicated that miR-23b-3p was upregulated in cartilage tissues of a patient with OA. However, the mechanism by which miR-23b-3p regulates the occurrence and development of OA remains unclear. Thus, this study aimed to investigate the role of miR-23b-3p in the progression of OA. METHODS: In this study, qRT-PCR was used to measure the expression of miR-23b-3p in OA tissue samples and normal controls, respectively. Western blotting assay was performed to detect the levels of collagen II, aggrecan, Bax and active caspase 3 in CHON-001 cells. In addition, the dual-luciferase reporter system assay was used to detect the interaction between miR-23b-3p and COL11A2 in OA. RESULTS: The levels of miR-23b-3p were upregulated, while the expressions of collagen II and aggrecan were decreased in OA tissues and in IL-1ß-treated CHON-001 cells. In addition, IL-1ß significantly induced apoptosis of CHON-001 cells via increasing the levels of Bax and active caspase 3. However, downregulation of miR-23b-3p markedly inhibited IL-1ß-induced apoptosis in CHON-001 cells via increasing the collagen II and aggrecan levels and decreasing Bax and active caspase 3 expressions. Meanwhile, dual-luciferase assay showed that COL11A2 was the direct target of miR-23b-3p in CHON-001 cells. Overexpression of miR-23b-3p markedly decreased the level of COL11A2 in cells. Moreover, downregulation of miR-23b-3p alleviated synovitis/cartilage destruction and reduced Osteoarthritis Research Society International scores and subchondral bone thickness in vivo. CONCLUSION: Downregulation of miR-23b-3p could alleviate the progression of OA through upregulating COL11A2 in vivo and in vitro. Therefore, downregulation of miR-23b-3p might be a potential therapeutic strategy for the treatment of OA.

Disocin prevents postmenopausal atherosclerosis in ovariectomized LDLR-/- mice through a PGC-1α/ERα pathway leading to promotion of autophagy and inhibition of oxidative stress, inflammation and apoptosis.

Atherosclerosis (AS) is one of the major causes leading to mortality of dysfunctional cardiovascular events in the menopausal women, which has long-term deficiency of estrogen. At present, the primary treatment for postmenopausal AS is hormone replacement therapy (HRT). However, it can increase the risks of ovarian and uterine cancers with long-term therapy. So seeking for a phytoestrogen which can overcome the disadvantages of HRT is a great mission. Dioscin, a traditional Chinese medicine, extracted from the roots of dioscorea nipponica, has anti-inflammatory, anti-tumor and anti-apoptosis activities. Especially, it also has estrogenic activity. Thus, this study aims to investigate the effects of dioscin on postmenopausal AS. Currently, ovariectomy (OVX) is the accepted model for AS associated with estrogen deficiency, and it can mimic the cessation of ovarian function that occurs in postmenopausal women as well. We used the high fat diet and ovariectomy(HFD-OVX)model to induce postmenopausal AS in the low-density lipoprotein receptor- deficient (LDLR-/-) mice. (1) The levels of TG, TC, LDL-C, HDLC, MDA, GSH, MDA and GSH in serum of HFD-OVX induced LDLR-/- mice were measured by colorimetric assay. (2) The artery injury of HFD-OVX induced LDLR-/- mice was detected with Oil Red O staining. (3) The protein expressions of NOX4, P22phox, IκB, p-p65, n-p65, ICAM-1, VCAM-1, caspase-3, caspase-9, bcl-2, PGC-1α, ERα, ERß in the arterial tissue of HFD-OVX induced LDLR-/- mice were detected by Western blot analysis. In vitro, the model of human aortic endothelial cells (HAECs) induced by oxidized low-density lipoprotein (ox-LDL) (150 µg /ml) was established, and the molecular mechanism of dioscin on atherosclerosis in postmenopausal women was investigated. (1) The levels of MDA, GSH, MDA and GSH in ox-LDL induced HAECs were measured by colorimetric assay. (2) Reactive Oxygen Species (ROS) of ox-LDL induced HAEC cells was detected by fluorescence staining. (3) The protein expressions of PGC-1α, ERα, ERß, NOX4, P22phox, IκB, p-p65, n-p65, ICAM-1, VCAM-1, caspase-3, caspase-9, bcl-2 and LC3 in ox-LDL induced HAECs were detected by Western blot analysis. (4) The autophagy level of ox-LDL induced HAECs was measured by transmission electron microscopy. (5) The applications of si-RNA transfection were used to explore whether dioscin could activate PGC-1α/ERα pathway to inhibit postmenopausal atherosclerosis. In vivo, we found that dioscin decreased the level of TG, TC, LDL-C and increased the level of HDLC in serum of HFD-OVX induced LDLR-/- mice, and it has protective effects to maintain the lipid homeostasis; The Oil Red O staining study showed that dioscin could significantly inhibit the formation of atherosclerotic plaques in HFD-OVX-treated LDLR-/- mice; Dioscin decreased the levels of NOX4, P22phox, p-p65, n-p65, ICAM-1, VCAM-1, caspase-3, caspase-9, but increased the levels of HDL-C, GSH, SOD, PGC-1α, ERα, ERß, IκB, Bcl-2 and elevated the autophagy level in arterial tissues of HFD-OVX induced LDLR-/- mice. It is particularly worth mentioning that the up-regulating effect of dioscin on ERα is stronger than ERß in OVX treated mice. In vitro, the results of colorimetric assay showed that dioscin decreased the level of MDA and LDH, increased the level of SOD and GSH in ox-LDL-induced HAEC cells; Dioscin also suppressed the release of ROS in ox-LDL-induced HAECs by fluorescence staining; Dioscin decreased the levels of NOX4, P22phox, p-p65, n-p65, ICAM-1, VCAM-1, caspase-3, caspase-9, but increased the levels of PGC-1α, ERα, ERß, IκB, Bcl-2 and the ratio of LC3-II/LC3-I in ox-LDL-induced HAECs; Dioscin significantly elevated the autophagy level of ox-LDL-induced HAECs by transmission electron microscopy analysis; In addition, by si-RNA transfection, we found that the inhibitory effects of dioscin on oxidative stress, inflammatory response and apoptosis might partly through PGC-1α/ERα pathway in ox-LDL induced HAECs. The data of dual-Luciferase reporter assay revealed that dioscin activated ERα at least partly through PGC-1α pathway. Dioscin significantly inhibited oxidative stress, inflammatory response, apoptosis and increased the level of autophagy in vivo and vitro. In addition, dioscin could regulate the balance of lipid metabolism. Moreover, we proved that the effects of dioscin attenuating postmenopausal atherosclerosis by inhibiting oxidative stress, inflammation and apoptosis were partly dependent on PGC-1α/ERα pathway. Therefore, dioscin, as a phytoestrogen, might become a drug for the treatment of atherosclerosis in postmenopausal women.

Up-regulated HIF-2α contributes to the Osteoarthritis development through mediating the primary cilia loss.

BACKGROUNDS: Up-regulated HIF-2α (hypoxia induced factor 2) had been demonstrated to contribute to Osteoarthritis (OA) development via inducing the expression of matrix-degrading enzymes. However, the HIF-2α also could promote primary cilia loss through HIF-2α/AURKA (Aurora kinase A)/NEDD9 pathway. And the primary cilia dysfunction is another characteristic of the OA. Thus, we investigated here whether the HIF-2α also contributes the OA development through mediating the primary cilia loss. METHODS: The primary chondrocytes were isolated from the experimental OA mice induced by destabilization of the medial meniscus (DMM). Chondrocytes were cultured under normoxia (21% O2) or hypoxia (2% O2) conditions. The HIF-1α and HIF-2α expressions were assessed by western blot. The cilia formation was counted by immuno-staining the acetylated tubulin. The contribution of HIF-1α or HIF-2α to the primary cilia loss was assessed by knocking-down the HIF-1α or HIF-2α individually. The HIF-2α/AURKA/NEDD9 pathway was validated through over-expressing or knocking-down specific components of the pathway and then counting the primary cilia number. Finally, the pathway was further confirmed in the OA mice. RESULTS: Hypoxia could induce the expression of both HIF-1α and HIF-2α, and also reduce the number of primary cilia on the chondrocytes isolated from the experimental OA mice. Knocking-down or over-expressing HIF-1α or HIF-2α individually showed that the HIF-2α could induce the primary cilia reduction rather than the HIF-1α. Manipulating the HIF-2α expression could positively affect the AURKA and NEDD9 expression. Manipulating the AURKA and NEDD9 expressions could reverse the function of HIF-2α on primary cilia. In the mice, knocking-down both AURKA and NEDD9 could alleviate the OA development significantly. CONCLUSION: Up-regulated HIF-2α contributes to the Osteoarthritis development through mediating the primary cilia loss, which might be developed as therapeutic targets for OA treatment.

A LINC00341-mediated regulatory pathway supports chondrocyte survival and may prevent osteoarthritis progression.

Osteoarthritis (OA) is the most common degenerative joint disease and results from progressive loss and destruction of articular cartilage and the underlying bone. The disease affects millions of people worldwide with an associated risk of mobility disability. However, the molecular basis underlying OA initiation and progression is not well understood and, currently, there is no effective intervention available to decelerate disease progression or restore degraded cartilage. We have found that lncRNA long intergenic nonprotein coding RNA 341 (LINC00341) is aberrantly downregulated in OA patient tissues and cultured OA chondrocytes. This is likely responsible for the increased apoptosis of chondrocytes and pathological destruction of cartilage. Further investigation has revealed that LINC00341 interacts with miR-141 to suppress its functional binding to the 3'-untranslated region of YY1-associated factor 2 (YAF2) messenger RNA. Aberrant downregulation of LINC00341 thus may ultimately lead to inhibition of the YAF2 protein, which has been implicated to be an antiapoptotic factor. Our study has revealed a new noncoding RNA-mediated regulatory network that highly likely protects chondrocytes by preventing apoptosis under normal conditions. The results will help further explore the molecular details pertaining to the progression of OA and stimulate efforts to develop effective therapies.

Study on the mechanism of excessive apoptosis of nucleus pulposus cells induced by shRNA-Piezo1 under abnormal mechanical stretch stress.

OBJECTIVE: The aim of the study was to explore the mechanism of excessive apoptosis of nucleus pulposus cells induced by short hairpin RNA (shRNA) Piezo type mechanosensitive ion channel component 1 (Piezo1) under abnormal mechanical stretch stress. METHODS: In vitro mechanical stretch stress model of nucleus pulposus cells in vitro was established, in which the expression of Piezo1 was interfered by transfection of shRNA-Piezo1 interfering vector. Both messenger RNA and protein level of Piezo1 were measured by reverse-transcription polymerase chain reaction and Western blot analysis, respectively. Cytoplasmic Ca2+ was detected by Fluo3-AM kit, and changes of mitochondrial membrane potential in cells were detected using Cell Meter Assay kit. Finally, the apoptosis was evaluated with annexin V-fluorescein isothiocyanate kit. RESULTS: The highest transfection efficiency of lentivirus titer was 1 × 10 TU/mL and the nucleus pulposus cells were transfected with plural multiplicity of infection = 50. Homo-3201 sequence exhibited the most effective silencing effect and was used in subsequent experiments as the default sequence of shRNA-Piezo1. The calcium content in the cytoplasm of the tension stress group increased significantly compared with that in the blank control group ( q = 3.773; P < 0.05). The level of cytosolic calcium in shRNA-interference group was significantly lower than that in stretch stress group ( q = 5.159; P < 0.05). Stretch stress treatment resulted in an elevated ratio of mitochondrial membrane potential turnover as opposed to blank control group ( q = 4.332; P < 0.05), while shRNA-interference group showed smaller ratio of mitochondrial membrane potential turnover than that in stretch stress group ( q = 4.974; P < 0.05). Similar results were also observed in apoptosis rate analysis ( q = 3.175; P < 0.05). CONCLUSION: ShRNA-Piezo1 can protect cells by reducing the level of intracellular Ca2+ and the change of mitochondrial membrane potential.

Luteolin attenuates glucocorticoid-induced osteoporosis by regulating ERK/Lrp-5/GSK-3ß signaling pathway in vivo and in vitro.

Glucocorticoid-induced osteoporosis (GIO) is a secondary osteoporosis with extensive use of glucocorticoids (GCs). GCs can increase bone fragility and fracture via inhibiting osteoblastic proliferation and differentiation. Luteolin (LUT), a kind of plant flavonoid, has been reported to exhibit the antioxidant activity, but the effects of LUT on GIO still remain unclear. This study aimed to investigate the effects of LUT on GIO both in vivo and in vitro and elaborate the potential molecular mechanisms. LUT increased the superoxide dismutase activity, glutathione level and decreased reactive oxygen species (ROS) level and lactate dehydrogenase release in GIO. Meanwhile, LUT decreased caspase-3, caspase-9, and Bax protein expressions and increased Bcl-2 protein expression in GIO. LUT increased the ratio of osteoprotegerin (OPG)/receptor activator of nuclear factor-κB Ligand (RANKL) messenger RNA (mRNA) expression and mRNA expression levels of osteogenic markers, including runt-related transcription factor 2, osterix, collagen type I, and osteocalcin. LUT also enhanced the extracellular signal-regulated kinases (ERK) phosphorylation, glycogen synthase kinase 3ß (GSK-3ß) phosphorylation, mRNA expression levels of lipoprotein-receptor-related protein 5 (Lrp-5) and ß-catenin. Further study revealed that Lrp-5 small interfering RNA (siRNA )and ERK-siRNA reduced the effects of LUT on GSK-3ß phosphorylation, alkaline phosphatase (ALP) activity and the ratio of OPG/RANKL mRNA expression. Moreover, ERK-siRNA decreased Lrp-5 mRNA expression in vitro. These results indicated that LUT promoted proliferation by attenuating oxidative stress and promoted osteoblastic differentiation by regulating the ERK/Lrp-5/GSK-3ß pathway in GIO. This study may bring to light the possible mechanisms involved in the action of LUT in GIO treatment, and benefit for further research on GIO.

Activating the PGC-1α/TERT Pathway by Catalpol Ameliorates Atherosclerosis via Modulating ROS Production, DNA Damage, and Telomere Function: Implications on Mitochondria and Telomere Link.

Catalpol, an iridoid glucoside, has been found present in large quantities in the root of Rehmannia glutinosa L. and showed a strong antioxidant capacity in the previous study. In the present work, the protective effect of catalpol against AS via inhibiting oxidative stress, DNA damage, and telomere shortening was found in LDLr-/- mice. This study also shows that activation of the peroxisome proliferator-activated receptor-γ coactivator-1α (PGC-1α)/telomerase reverse transcriptase (TERT) pathway, which is the new link between mitochondria and telomere, was involved in the protective effects of catalpol. Further, by using PGC-1α or TERT siRNA in oxLDL-treated macrophages, it is proved that catalpol reduced oxidative stress, telomere function, and related DNA damage at least partly through activating the PGC-1α/TERT pathway. Moreover, dual luciferase activity assay-validated catalpol directly enhanced PGC-1α promoter activity. In conclusion, our study revealed that the PGC-1α/TERT pathway might be a possible therapeutic target in AS and catalpol has highly favorable characteristics for the treatment of AS via modulating this pathway.

Arthroscopic debridement of anterior ankle impingement in patients with chronic lateral ankle instability.

BACKGROUND: The aim of this study was to determine the functional and radiological outcomes of arthroscopic treatment of anterior ankle impingement (AAI) in patients with chronic lateral ankle instability (CAI). METHODS: All patients with CAI between June 2012 and May 2015 were invited to participate in this investigation. All of them accepted open modified Broström repair of lateral ankle ligaments and were divided into two groups: AAI group (with anterior ankle impingement) and pure CAI group (without anterior ankle impingement). All of them were followed up using American Orthopaedic Foot and Ankle Society Score (AOFAS), Karlsson Ankle Functional Score and Tegner activity score. Ankle dorsiflexion was also examined. X-ray examination was applied to investigate anterior tibiotalar osteophytes. RESULTS: Finally, a total of 60 patients were followed up at a mean of 37 ± 10 months, including 22 patients in the AAI group and 38 patients in the pure CAI group. Preoperatively, the AAI group had significant lower AOFAS score (62.9 ± 11.7 vs 72.9 ± 11.1; p = 0.002) and Tegner activity score (1.5 ± 0.8 vs 2.1 ± 1.0; p = 0.04) respectively when compared with the pure CAI group. The ankle dorsiflexion of the AAI group (13 ± 2.1) was also significantly lower than that of the pure CAI group (26.2 ± 2.1) (p = 0.001). However, there was no significant difference in the AOFAS score or the Karlsson score or the Tegner score or the Ankle dorsiflexion between the two groups postoperatively. The postoperative X-ray images demonstrated complete osteophyte resection in all patients, and no recurrence of osteophyte. CONCLUSION: The functional outcome scores and dorsiflexion had significantly improved postoperatively. Combined treatment of chronic ankle instability and anterior ankle impingement produced satisfactory surgical outcomes in patients with CAI accompanied by anterior ankle impingement symptom.

[Expression characteristics of Piezo1 protein in stress models of human degenerative chondrocytes].

OBJECTIVE: To explore the expression characteristics of new mechanosensitive ion channel Piezo1 protein in stress models of human degenerative chondrocytes. METHODS: The stress stimulation model of human degenerative chondrocytes in vitro was constructed. Multi-channel cell stretch stress loading system FX-4000T was used to treat chondrocytes. According to the results of pre-test, the loading frequency of 0.5 Hz and the cell elongation of 20% were loaded. According to cell processing time, it was divided into 0 h, 2 h, 12 h, 24 h and 48 h mechanical stress group. The RT-PCR and Western-blot were used to test the expression of the Piezo1, also the Laser scanning confocal microscope (LSCM) was used to test the intensity of the fluorescence of the Piezo1. RESULTS: (1)The result of the RT-PCR showed that the expression of the Piezo1 in the 2 h group was higher than the 0 h group(F=13.917, q=0.037 1, P<0.05). The expression of the piezo1 in the 24 h group was the highest. While the expression of the piezo1 in the 48 h group was lower than the expression of the piezo1 in the 24 h group(F=13.917, q=0.049 5, P<0.05). (2)The result of the Western-blot showed that the 2 h group was higher than the 0 h group(F=19.341, q=0.037 1, P<0.05). The expression of the 24 h had the highest expression which was higher than the 48 h group(F=19.341, q=0.017 7, P<0.05). (3)The Piezo1 protein was extensively expressed in the cytoplasm and nucleus of the nucleus pulposus cells. And with the increase of stress processing time, the fluorescence intensity of the protein also increased. CONCLUSIONS: In human degeneration cartilage cells, the new mechanio sensitive ion channel Piezo1 protein has a trace expression. After loading periodic mechanical tensile force, the expression of Piezo1 protein increases with time dependence.