Mechanical unloading that results from aging, prolonged bed rest, or spaceflight leads to disuse osteoporosis (DOP), a significant public health concern. However, the mechanisms underlying this condition, as well as effective therapeutic strategies, remain incompletely defined. In this study, significant bone deterioration, increased osteoclast activation, and upregulation of the PI3K/AKT/NF-κB signaling cascade were observed in femoral specimens from hindlimb suspension (HLS) mice, a well-established DOP model. Phloretin, a naturally occurring flavonoid with notable antioxidant and anti-inflammatory activities, has shown anti-osteoclastogenic effects in chemically induced models. Nevertheless, its therapeutic efficacy and direct molecular targets in mechanical unloading-induced bone loss, a condition characterized by a distinct pathophysiology, remain largely undefined. Herein, we demonstrated that phloretin markedly suppressed osteoclastogenesis, attenuated excessive bone resorption and effectively counteracted bone loss in the hindlimb-unloading murine model. Complementing these in vivo findings, primary osteoclast cultures revealed that phloretin reduced osteoclast formation and activity by inhibiting the PI3K/AKT/NF-κB signaling axis. Importantly, pharmacological activation of the PI3K pathway with 740 Y-P significantly reduced Phloretin's inhibitory effects on osteoclast differentiation and bone protection, confirming that this pathway is the primary regulatory target. Unlike general anti-resorptive agents, phloretin specifically addresses the PI3K-mediated signaling activation triggered by mechanical unloading. Our findings provide robust evidence for the application of phloretin as a targeted therapy for disuse-related skeletal disorders.

The dynamic production and clearance of senescent osteoblasts affects bone homeostasis and health. However, the relationship between senescent osteoblasts and the immune system remains unclear. Here, a landscape of the interaction between immune cells and osteoblasts through spatial analysis of the bone microenvironment is presented. Sirtuin 1 (SIRT1), a longevity gene, regulates bone mass maintenance through a mechanism involving osteoblast-CD4+ cytotoxic T lymphocyte (CTL) crosstalk. In the osteoblastic niche, SIRT1 promotes the secretion of crucial chemokines, such as C-C motif chemokine ligand 3 (CCL3), C-C motif chemokine ligand 5 (CCL5), and C-X-C motif chemokine ligand 10 (CXCL10), by upregulating dishevelled-associated activator of morphogenesis 2 (DAAM2) through the acetylation of enhancer of zeste homolog 2 (EZH2), activating and recruiting CD4+ CTLs that eliminate senescent osteoblasts in a major histocompatibility complex class II (MHC-II)-dependent manner, slowing the bone ageing process and ameliorating osteoporosis. DAAM2 serves as a pivotal downstream effector for SIRT1 to exert immune-regulatory effects in the bone microenvironment; thus, targeting DAAM2 can treat osteoporosis by increasing CD4+ CTL responses. These results will facilitate the development of customised therapies targeting senescent osteoblasts to maintain bone health.

OBJECTIVE:To compare the clinical outcomes between group A (hybrid endoscopic technique: percutaneous uniportal endoscopic decompression combined with biportal endoscopic lumbar interbody fusion [LIF]) and group B (minimally invasive Quadrant transforaminal LIF) for lumbar spinal stenosis. METHODS:This study included 68 consecutive patients (group A: 32 patients; group B: 36 patients) retrospectively enrolled and treated between June 2019 and June 2021. Perioperative data were prospectively collected, including radiological outcomes (intervertebral disc height, sagittal Cobb angle, lumbar lordosis) and clinical outcomes (Oswestry Disability Index, visual analog scale [VAS], and Short Form 36 Health Survey [SF-36]). Paravertebral muscle injury was assessed by serum creatine phosphokinase levels. Additionally, fusion rate and perioperative complication rates were compared between the two groups. RESULTS:At 1 month postoperatively, group A demonstrated significantly better outcomes in VAS-Back, VAS-Leg, and SF-36 scores compared to group B (P < 0.05). At 3 months postoperatively, group A demonstrated significantly better outcomes in VAS-Back, Oswestry Disability Index, and SF-36 scores compared to group B (P < 0.05). Creatine phosphokinase levels in group A were significantly lower than those in group B at 1, 3, and 7 days postoperatively (P < 0.05). However, no significant intergroup differences were observed in intervertebral disc height, sagittal Cobb angle, and lumbar lordosis at 1 and 12 months postoperatively (P > 0.1). Additionally, fusion rates and surgical complication rates did not differ significantly between groups (P > 0.05). CONCLUSIONS:The hybrid endoscopic technique (percutaneous uniportal endoscopic decompression combined with biportal endoscopic LIF) demonstrates comparable clinical outcomes and fusion rates to minimally invasive Quadrant transforaminal LIF for lumbar spinal stenosis, while achieving reduced postoperative pain and improved early functional recovery.

Objective:To explore decompression strategies for lateral lumbar spinal stenosis under unilateral biportal endoscopy (UBE) assistance. Methods:A clinical data of 86 patients with lateral lumbar stenosis treated with UBE-assisted intervertebral decompression between September 2022 and December 2023 was retrospectively analyzed. There were 42 males and 44 females with an average age of 63.6 years (range, 45-79 years). The disease duration ranged from 6 to 14 months (mean, 8.5 months). Surgical levels included L 2, 3 in 3 cases, L 3, 4 in 26 cases, L 4, 5 in 42 cases, and L 5, S 1 in 15 cases. According to Lee's grading system, there were 21 cases of grade 1, 37 cases of grade 2, and 28 cases of grade 3 for lumbar spinal stenosis. Based on the location of stenosis and clinical symptoms, the 33 cases underwent interlaminar approach, 7 cases underwent interlaminar approach with auxiliary third incision, 26 cases underwent contralateral inclinatory approach, and 20 cases underwent paraspinal approach; then, the corresponding decompression procedures were performed. Visual analogue scale (VAS) score was used to evaluate lower back/leg pain before operation and at 1 and 3 months after operation, while Oswestry disability index (ODI) was used to evaluate spinal function. At 3 months after operation, the effectiveness was evaluated using the modified MacNab evaluation criteria. The spinal stenosis and decompression were evaluated based on Lee's grading system using lumbar MRI before operation and at 3 months after operation. Results:All procedures were successfully completed with mean operation time of 95.1 minutes (range, 57-166 minutes). Dural tears occurred in 2 cases treated with interlaminar approach with auxiliary third incision. All incisions healed by first intention. All patients were followed up 3-10 months (mean, 5.9 months). The clinical symptoms of the patients relieved to varying degrees. The VAS scores and ODI of lower back and leg pain at 1 and 3 months after operation significantly improved compared to preoperative levels ( P<0.05), and the indicators at 3 months significantly improved than that at 1 month ( P<0.05). According to the modified MacNab evaluation criteria, the effectiveness at 3 months after operation was rated as excellent in 52 cases, good in 21 cases, and poor in 13 cases, with an excellent and good rate of 84.9%. No lumbar instability was detected on flexion-extension X-ray films during follow-up. The Lee's grading of lateral lumbar stenosis at 2 days after operation showed significant improvement compared to preoperative grading ( P<0.05). Conclusion:For lateral lumbar spinal stenosis, UBE-assisted decompression of the spinal canal requires the selection of interlaminar approach, interlaminar approach with auxiliary third incision, contralateral inclinatory approach, and paraspinal approach based on preoperative imaging findings and clinical symptoms to achieve better effectiveness.

Introduction This study examines bone turnover marker (BTM) variations between bone marrow and peripheral blood in osteoporotic and non-osteoporotic patients. BTMs offer insights into bone remodeling, crucial for understanding osteoporosis. Methods A total of 133 patients were categorized into osteoporotic and non-osteoporotic cohorts. BTMs—C-telopeptide cross-linked type 1 collagen (β-CTX), serum osteocalcin (OC), Procollagen type I N-propeptide (P1NP), 25(OH)D—were measured in bone marrow and peripheral blood. Lumbar spine bone mineral density (BMD) was assessed. Results Osteoporotic patients exhibited elevated β-CTX and OC levels in peripheral blood, indicating heightened bone resorption and turnover. β-CTX levels in osteoporotic bone marrow were significantly higher. Negative correlations were found between peripheral blood β-CTX and OC levels and lumbar spine BMD, suggesting their potential as osteoporosis severity indicators. No such correlations were observed with bone marrow markers. When analyzing postmenopausal women separately, we obtained consistent results. Conclusions Elevated β-CTX and OC levels in osteoporotic peripheral blood highlight their diagnostic significance. Negative β-CTX and OC-BMD correlations underscore their potential for assessing osteoporosis severity. Discrepancies between peripheral blood and bone marrow markers emphasize the need for further exploration. This research advances our understanding of BTM clinical applications in osteoporosis diagnosis and treatment.

Abstract During ageing, the dynamic balance between the production and clearance of senescent osteoblasts affects bone homeostasis and health. However, the relationship between senescent osteoblasts and the immune system remains unclear. Here, we provide evidence that SIRT1, known as a longevity gene, could orchestrate bone mass maintenance through a novel mechanism involving osteoblast- CD4+ cytotoxic T lymphocyte (CTL) crosstalk. In the osteoblastic niche, SIRT1 promotes the secretion of key chemokines (e.g. CCL3, CCL5, and CXCL10) by upregulating DAAM2 through acetylation of EZH2 protein, thereby activating and recruiting CD4+ CTLs. Then, CD4+ CTLs eliminate senescent osteoblasts in an MHC-II-dependent manner, slowing the bone ageing process and effectively alleviating osteoporosis. This mechanism shows that DAAM2 is a pivotal downstream effector for SIRT1 to exert immune-regulatory effects in the bone microenvironment, and that targeting DAAM2 could accurately treat osteoporosis by increasing CD4+ CTL responses. This work enables a multi-perspective assessment of physiological bone senescence, paving the way for customised therapies targeting senescent osteoblasts to maintain bone health.

Background and Objective: Spinal cord injury (SCI) is typically the result of compressive forces on spinal cord tissue, leading to loss of motor or sensory function. As a cheap and biocompatible substance,gamma-Oryzanol has been shown to cross the blood-brain barrier and exert beneficial effects on neural tissue. Our Previous studies have demonstrated the potential of a hydrogel based on the cross-linking of oxidized hyaluronic acid (OHA) and Hyaluronic Acid-Adipic Dihydrazide (HA-ADH) for treating osteoarthritis by loading Selenium Nanoparticles (SeNPs) through a Schiff base reaction. This study investigated the use of this hydrogel loaded with gamma-Oryzanol to treat SCI in mice. Materials and Methods: Synthesize the hydrogel according to the previous method and load the appropriate concentration of gamma-Oryzanol. Mice were randomly divided into groups and treated accordingly. Within 6 weeks after surgery, we used multiple methods including Basso mouse scale (BMS) score to conduct behavioral tests and analyzed the lesion area by Hematoxylin and Eosin (H&E) staining and immunofluorescence staining. Results: The study showed that mice treated with gamma-Oryzanol showed better motor function. Observation of the lesion area showed that the inflammatory and demyelinating conditions of the injury site in the mice treated with gamma-Oryzanol improved and more axons related to motor function were preserved with less scar formation. Endoplasmic reticulum stress-related proteins in the spinal cord tissue of gamma-Oryzanol-treated mice decrease. Conclusion: Although further research is warranted, we propose that gamma-Oryzanol, as a relatively inexpensive dietary phytochemical, has potential clinical value.

Spinal cord injury (SCI) usually arises from compression due to traffic accidents and falls, resulting in varying degrees of movement, sensory loss, and possible paralysis. Glabridin (Gla) is a natural compound derived from licorice. It significantly affects drug development and medicine because of its anti-inflammatory, anti-oxidative, anti-tumoral, antibacterial, bone protective, cardiovascular protective, neuroprotective, liver protective, anti-obesity, and anti-diabetic properties. Various methods were employed to administer Gla to SCI mice in order to investigate its impact on the recovery of motor function. The mice were allocated into four cohorts using a randomization procedure. In the sham cohort, solely the lamina of vertebral arch was surgically exposed without causing any harm to the spinal cord tissue. Conversely, the injury cohort was subjected to spinal cord tissue damage and received no treatment thereafter. The mice in the remaining two cohorts received a dosage of 40 mg/kg Gla every two days via either intraperitoneal or intrathecal injection for a duration of 42 d following spinal cord injury. We conducted behavioral tests utilizing the Basso Mouse Scale score and gait analysis techniques. Magnetic resonance imaging and hematoxylin and eosin were employed to evaluate scar tissue formation. Systemic inflammation in mice was evaluated by employing an enzyme-linked immunosorbent assay. Gla promoted motor function recovery in mice following SCI and improved the pathological environment in the damaged area. These alterations were more evident in mice subjected to the intrathecal injection method. Intraperitoneal injections appear to be more beneficial for controlling systemic inflammatory responses. Although more intensive studies are required, Gla exhibits promising clinical potential as a cost-effective dietary phytochemical.

Osteoporosis is a serious systemic metabolic bone system disease.This study aimed to identify the target genes of isopsoralen and the signaling pathways involved in the differential expression of the genes involved in osteoclast differentiation. We hypothesized that isopsoralen may inhibit osteoclast differentiation by blocking the nuclear factor kappa-B (NF-κB) signaling pathway and verified our hypothesis through basic experiments. The 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay was used to detect the effect of isopsoralen on the proliferation and viability of primary mouse bone marrow monocytes (BMMCs). The effect of isopsoralen on receptor activator of nuclear factor kappa-B ligand (RANKL)-induced osteoclast differentiation was determined by using tartrate-resistant acid phosphatase (TRAP) staining. Quantitative real-time PCR (qRT-PCR) and Western blot were used to detect the expression of the related genes and proteins. The Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway of isopsoralen target genes were obtained through comprehensive analysis using the STITCH database, Cytoscape 3.8.2, and R-Studio software. Differentially expressed genes (DEGs) were found in osteoclasts induced by RANKL before and after 3 days using R-Studio, following which KEGG analysis was performed. Next, enrichment analysis was performed on the KEGG pathway shared by the target genes of isopsoralen and the differentially expressed genes during osteoclast differentiation to predict the signaling pathway underlying the inhibition of osteoclast differentiation by isopsoralen. Finally, Western blot was used to detect the effect of isopsoralen on the activation of signaling pathways to verify the results of our bioinformatics analysis. Based on the enrichment analysis of isopsoralen target genes and differentially expressed genes during osteoclastogenesis, we believe that isopsoralen can inhibit RANKL-induced osteoclastogenesis by inhibiting the NF-κB signaling pathway.

Objective: Spinal cord injury (SCI) is usually caused by compression of spinal cord tissue. The main manifestations are the loss of motor or sensory functions. Because injured individuals are usually relatively young and have no effective treatment measures, there is a heavy economic burden on their families and society. Rice was the most abundant source of GORZ. This compound exhibits both antioxidant and anti-inflammatory properties. gamma-Ory-zanol has been shown to cross the blood-brain barrier intact and benefit brain function. This study is the first time the effect of gamma-Oryzanol on the recovery of motor function after SCI in mice. Therefore, we injected gamma-Oryzanol into SCI mice in different ways to observe the effect on motor function recovery of mice.Materials and methods: The mice were randomly divided into 4 groups. In one group, only lamina was opened without damaging the spinal cord tissue (sham group); One group was left untreated after the injured spinal cord tissue (injured group). In the other two groups, mice were given 100 mg/kg of gamma-Oryzanol every 2 days through the intraperitoneal or intrathecal for 42 days after injuring the spinal cord. Behavioral tests were performed using the Basso Mouse Scale (BMS) scores and gait analysis. Hematoxylin and Eosin (H&E) and immunofluorescence staining were used to observe changes in the lesion area, demyelination, axonal regeneration, and scar tissue formation.Results: 1. Behavioral experiments showed that the gamma-Oryzanol-treated mice showed better exercise capacity than those in the injured group. 2. The pathology and demyelination at the injury site were improved in gamma-Oryzanol-treated mice. 3. gamma-Oryzanol-treated mice had more M2-type macrophages at the site of injury. At the same time, gamma-Oryzanol-treated mice retained more axons associated with motor function and less scarring.Conclusions: Studies have shown that gamma-Oryzanol can promote the recovery of motor function in mice after SCI. And the change was even more pronounced in mice treated with intrathecal injection methods. Although more intensive studies are needed, we suggest that gamma-Oryzanol has potential clinical value as a recognized inexpensive dietary phytochemical.

Currently, resistance to oxaliplatin (OXA) has become an important obstacle to improving the clinical outcome of patients with colorectal cancer (CRC). Moreover, long non-coding RNAs (lncRNAs) have been documented in cancer chemoresistance, and our bioinformatic analysis suggested an involvement of lncRNA CCAT1 in CRC development. In this context, this study aimed to clarify the upstream and downstream mechanisms underpinning the effect of CCAT1 in the resistance of CRC to OXA. The expression of CCAT1 and the upstream B-MYB in the CRC samples was predicted by bioinformatics analysis and then verified using RT-qPCR in CRC cell lines. Accordingly, overexpression of B-MYB and CCAT1 was observed in CRC cells. SW480 cell line was used for the construction of OXA-resistant cell line (SW480R). Ectopic expression and knockdown experiments of B-MYB and CCAT1 were conducted in SW480R cells to delineate their roles in the malignant phenotypes and half-maximal (50%) inhibitory concentration (IC50) of OXA. It was found that CCAT1 promoted the resistance of CRC cells to OXA. Mechanistically, B-MYB transcriptionally activated CCAT1, which recruited DNMT1 to inhibit SOCS3 expression through elevating the SOCS3 promoter methylation. By this mechanism, the resistance of CRC cells to OXA was enhanced. Meanwhile, these in vitro findings were reproduced in vivo on xenografts of SW480R cells in nude mice. To sum up, B-MYB might promote the chemoresistance of CRC cells to OXA via regulating the CCAT1/DNMT1/SOCS3 axis.

目的 探讨大蒜素是否具有抑制硬膜外疤痕来源的成纤维细胞增殖及迁移的作用.方法 培养人体硬膜外瘢痕来源的成纤维细胞至对数增长期,用不同浓度梯度大蒜素处理24 h.CCK-8法检测细胞活性、镜下观察细胞形态变化及划痕试验检测细胞迁移能力.结果 不同浓度大蒜素处理成纤维细胞24 h后,与对照组比较,实验组细胞活性降低,并呈浓度依赖性(P<0.05);显微镜下发现实验组细胞萎缩变形,体积变小,核固缩、碎裂;AO/EB染色后实验组细胞体积变小,膜皱缩,胞浆减少,核固缩,出现凋亡小体等凋亡特征,随着大蒜素浓度增加凋亡细胞及死细胞增多,与对照组比较,细胞迁移能力降低并具有浓度依赖性(P<0.05).结论 大蒜素能够呈浓度依赖性抑制硬膜外疤痕来源的成纤维细胞增殖及迁移.

背景:研究报道大蒜素具有抗纤维化作用,可抑制过度增生的成纤维细胞增殖和胶原合成能力,但大蒜素抑制成纤维细胞增殖及诱导其凋亡的确切机制尚未明确.目的:探讨大蒜素抑制硬膜外瘢痕来源的成纤维细胞增殖、迁移并诱导其凋亡的作用机制,为防治硬膜外粘连提供新思路.方法:培养人体硬膜外瘢痕来源的成纤维细胞至对数增长期,用不同质量浓度梯度大蒜素处理24 h.采用荧光显微镜观察细胞形态,CCK-8试验检测细胞增殖,划痕试验及Transwell试验检测细胞迁移能力,流式细胞术检测细胞凋亡,Western blot及实时定量PCR技术检测细胞增殖、凋亡相关蛋白和基因表达水平.结果与结论:①大蒜素处理24 h后,成纤维细胞萎缩变形,体积变小,细胞核固缩、碎裂;②细胞增殖能力和迁移率均呈剂量依赖性下降(P<0.01);③G1期细胞比例呈剂量依赖性升高(P<0.01),而S期和G2期呈剂量依赖性下降(P<0.01),流式细胞仪结果显示凋亡细胞比例呈剂量依赖性升高(P<0.01);④与对照组(未加入大蒜素处理组)相比,细胞核增殖抗原及Bcl-2在蛋白与mRNA水平的表达均显著降低(P<0.01),而Bax表达显著增高(P<0.01);⑤说明大蒜素具有抑制硬膜外瘢痕来源的成纤维细胞增殖、迁移并促其凋亡的作用,可能与调控细胞核增殖抗原、Bcl-2及Bax表达有关.

Vertebroplasty is the most widely used method for treating osteoporotic vertebral compression fractures (OVCF). During this procedure, bone cement is injected into the vertebral body. Fracture and additional fractures can occur adjacent to the treatment site. Thus, we studied factors causing such vertebral fractures after vertebroplasty and calculated the appropriate amount of bone cement to inject. From September 2012 to March 2016, 187 patients with OVCF undergoing vertebroplasty were selected, and 112 patients with complete follow-up information were selected. Of these, 28 had adjacent vertebral fractures (refracture group) during the follow-up period, and 84 patients had no adjacent vertebral fractures (control group). Then, sex, age, body weight, bone mineral density (BMD), and bone cement injection (bone cement injection volume and bone fracture vertebral volume percent) were compared. All patients had significant pain relief within 24 h (preoperative and postoperative [24 h later] VAS scores were 7.4 ± 0.8 and 2.3 ± 0.5, respectively). The age and weight were not statistically significantly different (P > 0.05). BMD values were statistically significantly different between groups as was sex (P < 0.05). Bone cement injection volume, BMD values, and sex were statistically significantly related to adjacent vertebral fractures after vertebroplasty, and cement injection volumes exceeding 40.5% caused adjacent vertebral fractures.

OBJECTIVE: We compared the mid-term efficacy and safety of anterior cervical discectomy and fusion (ACDF) using a Zero-Profile device for cervical degenerative disc disease (CDDD) with and without osteoporosis. METHODS: We performed a retrospective study of elderly patients with CDDD treated by single-level ACDF with a Zero-Profile device. The patients were divided into group A (osteoporosis) and group B (no osteoporosis) according to the bone mineral density. The clinical outcomes (Japanese Orthopaedic Association, neck disability index, visual analog scale, and short-form 36 scores), radiological outcomes (cervical lordosis and fusion rate), and complications were reviewed at each follow-up examination. RESULTS: All procedures were successfully performed in all patients. The Japanese Orthopaedic Association, neck disability index, visual analog scale, and short-form 36 scores and cervical lordosis were significantly improved postoperatively in both groups (P < 0.05). However, no significant difference was found between the 2 groups at each follow-up point (P > 0.05). No significant difference was found in the fusion rate at 3 months postoperatively (group A, 88.9%; group B, 90.0%), dysphagia rate at 1 month postoperatively (group A, 11.1%; group B, 15.0%), or cage subsidence rate at the final follow-up visit (group A, 11.1%; group B, 10.0%; P > 0.05). All patients achieved solid fusion, and no patient had dysphagia at the final follow-up examination. CONCLUSIONS: ACDF with the Zero-Profile device can be used as an effective and reliable treatment for single-level CDDD with osteoporosis.

The present study aimed to evaluate whether the application of tacrolimus (FK506) could improve functional recovery in spinal cord injury (SCI) rat models by activating astrocytes, and to further investigate the underlying mechanisms of this action. Male Sprague-Dawley rats (n=56) were used to establish moderate SCI models, which were induced at the T10 spinal segment by dropping a 10-g weight from a height of 25 mm using a New York University Impactor device. The rats were randomly separated into the FK506 or control group (n=28 per group). Rats were treated with FK506 (0.5 mg/kg) or saline intravenously 30 min after sustaining the injury. Functional recovery was evaluated over 42 days following the injury, and epidermal growth factor (EGF) levels were detected. The astrocytes were treated with FK506 in vitro, and the EGF mRNA and protein expression levels were analyzed using reverse transcription-quantitative polymerase chain reaction and ELISA, respectively. DNA microarray analysis was also performed to evaluate the genes in astrocytes. Rats in the FK506 group had improved locomotor functional recovery compared with those of control group. Furthermore, FK506 upregulated EGF expression of astrocytes both in vivo and in vitro. Subsequent to treatment with FK506-conditioned medium (CM), the length of neuronal cells increased 61.06% on the first day, and increased 56.4% on the third day compared with those of C-CM group. Furthermore, addition of anti-EGF neutralizing antibodies could interrupt the promotion of neurite outgrowth by FK506-CM. The present study indicates that astrocytes have an important role as mediators of FK506-improved spinal cord function recovery, and this partially clarifies the role of cell-cell interaction through modulating EGF in this process.

OBJECTIVE:To illustrate the role of epidermal growth factor (EGF) secreted by astrocytes in the process of tacrolimus (FK506) in promoting neurite outgrowth. METHODS:The spinal cord astrocytes and neuronal cells were isolated respectively from 2-day-old Sprague Dawley (SD) rats and 15-day SD pregnant rats, and cultured in vitro and identified by immunofluorescence staining. The spinal cord astrocytes were cultured with 20 µmol/L FK506 medium in the experimental group, and with FK506 free medium in the control group. The supernatant was collected after 24 hours for preparing conditioned medium, and astrocytes were collected. EGF proteins in the conditioned medium were detected with ELISA, and EGF gene expressions of astrocytes were detected with real-time quantitative PCR (RT-qPCR). The spinal cord neurons were cultured respectively with conditioned medium from the experimental group (FK506-CM) and the control group (C-CM) in group A and group B, also with neutralized C-CM and neutralized FK506-CM with anti-EGF neutralizing antibodies in group C and group D. Both the total neurite length and the longest neurite length were measured and compared among groups. RESULTS:Both astrocytes and neurons were confirmed by immunofluorescence staining. The EGF content of experimental group (0.241 ± 0.044) was significantly higher than that of the control group (0.166 ± 0.014) (t = 3.93, P = 0.01); EGF gene expression of the experimental group (1.12 ± 0.25) was significantly higher than that of the control group (0.46 ± 0.11) (t = 5.78, P = 0.00). The neurite length measurement displayed that the total neurite length and the longest neurite length of groups C and D were significantly shorter than those of groups A and B (P < 0.05). Both the total and longest neurite length of group A were significantly longer than those of group B (P < 0.05), but no significant difference was shown between groups C and D (P > 0.05). CONCLUSION:The EGF secreted by spinal cord astrocytes can promote the neurite outgrowth. So spinal cord astrocytes can be used as an important intermediary target of FK506 to promote the recovery of neurological function.

Mitomycin C (MMC) is known to reduce epidural fibrosis, but the underlying mechanisms have not yet been elucidated. Aberrant miR-200b expressions have been reported in multiple types of fibrotic tissues from many diseases. The aim of this study was to clarify the mechanism by which MMC induces fibroblasts apoptosis and reduces epidural fibrosis. The expression of miR-200b in human fibroblasts was determined after MMC treatment, and the targeted association between miR-200b and RhoE was determined using the luciferase activity assay. The effects of MMC and miR-200b on human fibroblasts apoptosis were evaluated using flow cytometry and western blot analysis. The effects of MMC and miR-200b on epidural fibrosis were evaluated using the Rydell classification, hydroxyproline content, apoptotic cell count and histological analysis. The study revealed that MMC could significantly downregulate miR-200b expression and induce human fibroblasts apoptosis. The direct downregulation of miR-200b could induce human fibroblasts apoptosis. Furthermore, we identified the binding sequence for miR-200b within the 3' untranslated region of RhoE. RhoE was confirmed to be a direct target of miR-200b, and RhoE itself acted as a promoter of fibroblasts apoptosis. The inhibition of miR-200b increased fibroblasts apoptosis and reduced epidural fibrosis in rats, which was in accordance with the effect of MMC. This study suggests that MMC induces fibroblasts apoptosis and reduces epidural fibrosis by regulating miR-200b expression and its targeting of RhoE.

The purpose of this study is to compare clinical outcome of different surgical methods in treating multi-segmental cervical degeneration. Three hundred and sixty eight patients with multi-segmental cervical degeneration were retrospectively selected and divided into two groups with 184 cases in each based on different surgical methods: one group accepted surgeries from anterior surgical approach and the other group accepted surgeries from posterior surgical approach. Perioperative parameters including operative time, intraoperative blood loss and length of stay were compared between two groups. Patients were followed up after 1 week, 6 month, 10 months and 1 year after surgery. Cervical X-ray was retaken, and Japanese orthopaedic association (JOA) scores, neck disability index (NDI ) scores and numerical pain rating scale (NPRS ) scores were obtained for comparison. Samples from cervical disc were processed to detect cytokines level including IL-1, IL-6, TNF-α and MMP-3. Perioperative parameters including operative time, intraoperative blood loss and length of stay showed no significant difference (P < 0.05) between the two groups. JOA score, NDI scores and NPRS scores, all showed a significant improvement after the surgery in both methods, however, when comparing the two methods, no significant difference was found between two groups (P > 0.05), except that NDI scores in anterior surgical approach group were significantly lower than posterior surgical approach group at different follow-up time points (P < 0.05). The average height of fused vertebral bodies after surgery in two groups was significantly different from pre-operative height (P < 0.05), and angle loss in posterior surgical approach group was significantly higher than anterior surgical approach (P < 0.05), which was statistically different. Cytokines including IL-1, IL-6, TNF-α and MMP-3 in two groups had no statistical difference (P > 0.05). Anterior approach surgery and posterior approach surgery are both effective methods to treat multi-segmental cervical degeneration. Anterior approach had better clinical outcomes within 1-year follow-up.

The objective of this study was to investigate the expression of adiponectin receptors (AdipoR1, R2, and T-cadherin) in both normal subjects and patients with knee osteoarthritis (OA). We used immunofluorescence to assess expression of adiponectin receptors in the chondrocytes of normal subjects (n = 3) and OA patients (n = 3). We also studied mRNA expression of adiponectin receptors in both groups by real-time polymerase chain reaction (real-time PCR). Finally, we utilized Western blotting to confirm the presence of adiponectin receptors. As compared with osteoarthritic chondrocytes, normal chondrocytes showed stronger immunoreactivity for AdipoR1, AdipoR2, and T-cadherin. The expression levels of both AdipoR1 and AdipoR2 mRNA were significantly lower in the osteoarthritic chondrocytes compared with those in the normal chondrocytes, 19 ± 2 and 36 ± 3 % of normal chondrocytes, respectively (P < 0.001). T-cadherin mRNA expression levels of the osteoarthritic chondrocytes were also lower than those in the normal chondrocytes, but not statistical significant (P = 0.072). The expression levels of AdipoR1 and AdipoR2 protein were significantly higher in the normal chondrocytes compared with those in the osteoarthritic chondrocytes (P < 0.001, P < 0.01, respectively). T-cadherin protein expression level of the normal chondrocytes was also higher than those in the osteoarthritic chondrocytes, but the difference is not statistical significant (P = 0.114). Expression of adiponectin receptors protein in normal and osteoarthritic chondrocytes is consistent with its mRNA expression levels. In conclusion, we report for the first time down-regulation of adiponectin receptors (AdipoR1, R2, and T-cadherin) in osteoarthritic chondrocytes. Decreased adiponectin receptors in OA may reduce the tissue sensitivity to adiponectin, thus lost the protection from adiponectin in the progression of OA.