1.Changes of upper airway in patient with skeletal class Ⅱ malocclusion accompanied by OSAHS after maxillomandibular advancement surgery:A case report and literature review
Xiangjin HU ; Xiumei SUN ; Kai CHEN ; Guomin WU
Journal of Jilin University(Medicine Edition) 2025;51(2):493-500
The patients with skeletal class Ⅱ malocclusion are often accompanied by structural and functional abnormalities of the upper airway,and obstructive sleep apnea hypopnea syndrome(OSAHS)may occur in severe cases.The morphologic and fluid dynamic changes of the upper airway in one patient with skeletal class Ⅱ malocclusion accompanied by OSAHS after maxillomandibular advancement(MMA)surgery were observed.The patient was a 27-year-old male with skeletal class Ⅱ malocclusion and moderate OSAHS.Combined orthodontic and orthognathic treatment was conducted,involving the anterior movement of the maxilla and mandible.After surgery,the patient's facial and intraoral features showed a satisfactory occlusion relationship with normal overbite,overjet and canine-molar relationships.There was also a significant improvement in the upper airway flow field.Two years after surgery,the cross-sectional area of the nasopharyngeal airway increased by 10.76%with a 55.36%reduction in pressure.The oropharyngeal airway showed a 108.25%increase in cross-sectional area with a 98.14%pressure reduction.The hypopharyngeal airway exhibited a 97.51%increase in cross-sectional area with a 351.03%pressure reduction.The laryngopharyngeal airway demonstrated a 27.54%increase in cross-sectional area with a 95.62%pressure reduction.The apnea-hypopnea index(AHI)decreased by 55.45%,achieving the treatment goal.Morphological measurements combined with fluid dynamic analysis can comprehensively evaluate the condition of the upper airway.The approximate value of the critical closing pressure(Pcrit)obtained from computational fluid dynamics(CFD)analysis may serve as a simple quantitative indicator for assessing the upper airway condition.
2.Anti-3-hydroxy-3-methylglutaryl-coenzyme A reductase antibody-mediated necrotizing myopathy in 10 children: clinical features and treatment outcomes
Qing WU ; Qianying LYU ; Haimei LIU ; Wanzhen GUAN ; Yinyu GONG ; Yifan LI ; Qiaoqian ZENG ; Xiaomei ZHANG ; Qijiao WEI ; Ling YANG ; Guomin LI ; Xihua LI ; Lei ZHAO ; Li SUN
Chinese Journal of Rheumatology 2025;29(7):575-582
Objective:To analyze the clinical characteristics and treatment outcomes of children with anti-3-hydroxy-3-methylglutaryl-coenzyme A reductase (anti-HMGCR) antibody-mediated necrotizing myopathy, and to explore early identification and management strategies to provide reference for clinical diagnosis and treatment.Methods:A retrospective analysis was conducted on the clinical data and treatment outcomes of 10 pediatric patients with anti-HMGCR antibody-mediated necrotizing myopathy admitted to the Department of Rheumatology, Children′s Hospital of Fudan University from December 2020 to December 2024. Statistical description was performed using SPSS 22.0.Results:Among the 10 patients, the male-to-female ratio was 1:4, the age of onset was (7.2±4.0) years, and the disease duration at diagnosis was (22.2±19.6) months. None had a history of statin exposure. Six patients presented with muscle weakness, and4 were diagnosed due to asymptomatic elevation of creatine kinase (CK); 4 had dermatomyositis-like rashes. All patients showed significantly elevated CK levels [median 3 291(1 969, 8 776)U/L] and underwent muscle biopsy. Histopathological findings revealed myofiber degeneration, necrosis, and regeneration in all cases, with inflammatory infiltration in 9 cases, MHC-Ⅰ positivity in all, and C5b-9 positivity in 9 cases. The median follow-up duration was (15.7±6.3) months. At the last follow-up, muscle strength was normal or nearly normal, and the CK median value had decreased to 977.5 (211.0, 3 536.0) U/L.Conclusion:For patients with suspected idiopathic inflammatory myopathy and significantly elevated CK, muscle-specific antibody testing-including anti-HMGCR-and muscle biopsy should be performed promptly regardless of the presence of skin rash, to ensure accurate diagnosis and guide treatment, thereby avoiding misdiagnosis or missed diagnosis.
3.Clinical characteristics of juvenile dermatomyositis in anti-nuclear matrix protein 2 antibody-positive patients and risk factors for severity: a national multicenter retrospective study
Huiyuan YANG ; Wanzhen GUAN ; Ling2 YANG ; Haimei LIU ; Xiaoqing3 LI ; Haiguo YU ; Meiping LU ; Jun YANG ; Xiaohui LIU ; Hongxia ZHANG ; Wei ZHANG ; Jihong XIAO ; Xiaozhong LI ; Guomin LI ; Hong CHANG ; Sheng HAO ; Yue DU ; Daliang XU ; Ling WU ; Wenjie ZHENG ; Li LIU ; Xinhui JIANG ; Shaohui ZHU ; Dongmei ZHAO ; Xuemei TANG ; Li SUN
Chinese Journal of Pediatrics 2025;63(12):1299-1305
Objective:To investigate the clinical characteristics and independent risk factors of severe disease in patients with anti-nuclear matrix protein (NXP) 2 antibody-positive juvenile dermatomyositis (JDM).Methods:A retrospective cohort study was conducted, including 219 anti-NXP2 antibody-positive JDM patients admitted to 23 children′s hospitals across China from July 2011 to July 2023. Patients were classified into severe and non-severe groups based on classification criteria for severe dermatomyositis. Demographic characteristics, clinical manifestations, and laboratory parameters were compared between the 2 groups using independent sample t-test, Mann-Whitney U test, or χ2 test. Univariate and multivariate Logistic regression analyses were performed to identify risk factors for severe disease. The receiver operating characteristic curve was employed to calculate optimal cut-off values. Results:Among the 219 patients, 108 were male and 111 were female, with an age at onset of 6.3 (3.5, 9.4) years. The severe group comprised 69 patients, and the non-severe group 150 patients. The severe group had significantly higher rates of fever, heliotrope rash, subcutaneous edema, periorbital edema, anti-Ro52 antibody positivity, as well as elevated levels of ferritin-to-albumin ratio (FAR), creatine kinase (CK), aspartate aminotransferase (AST), and lactate dehydrogenase (LDH) (all P<0.05). Multivariate analysis identified anti-Ro52 antibody positivity ( OR=13.26, 95% CI 1.37-128.29) and elevated FAR ( OR=1.90, 95% CI 1.09-2.31) as independent risk factors for severe anti-NXP2 antibody-positive JDM (both P<0.05). Receiver operating characteristic curve analysis revealed that a FAR cutoff value of 6.82 predicted severe disease with an area under the curve of 0.87 (95% CI 0.81-0.94, P<0.001), sensitivity of 0.85, and specificity of 0.70. All patients received glucocorticoid therapy, and the severe group received higher proportions of steroid pulse therapy, cyclophosphamide, mycophenolate mofetil, intravenous immunoglobulin, biologics, and adjuvant treatments compared to the non-severe group (all P<0.05). In terms of outcomes, 2 patients (2.9%) in the severe group died (due to neurological involvement and intestinal perforation, respectively), while the remaining patients achieved complete clinical response or remission. All patients in the non-severe group achieved remission. Conclusions:The primary clinical features of anti-NXP2 antibody-positive JDM included fever, heliotrope rash, subcutaneous edema, periorbital edema, anti-Ro52 antibody positivity, and elevated levels of CK, AST, LDH, and FAR. Furthermore, anti-Ro52 antibody positivity and a FAR>6.82 were identified as independent risk factors.
4.Exploration on the intervention mechanism of Zhuanggu Zhitong Capsules in postmenopausal osteoporosis based on JNK signaling molecules
Meihua WU ; Ronghui LI ; Yunfeng YU ; Bing GUO ; Guomin ZHANG ; Qinghu HE ; Xiaoming LEI ; Xinbin XIA
International Journal of Traditional Chinese Medicine 2025;47(5):630-637
Objective:To investigate the effects of Zhuanggu Zhitong Capsules on JNK signaling molecules and their phosphorylated proteins in postmenopausal osteoporosis model female mice.Methods:The rats were divided into sham-operation group, blank group, model group, positive drug group, and Zhuanggu Zhitong Capsules group according to the random number table method, with 10 rats in each group. The model group, the positive drug group and the Zhuanggu Zhitong Capsules group were prepared by bilateral ovarian detomy to prepare a female mouse model of postmenopausal osteoporosis. The positive drug group was given 0.9 mg/kg of alendronate sodium, the Zhuanggu Zhitong Capsules group was given was Zhuanggu Zhitong Capsules 1.944 g/kg for gavage, and the blank group, sham-operation group, and model group were given the same volume of normal saline for gavage, once a day for a total of 13 weeks. Rat vaginal exfoliated cells were stained with Wright's staining; serum Omentin-1 and 25(OH)D 3 levels were determined by ELISA; renal tissue and femoral structure were observed by HE staining; JNK and p-JNK protein expressions were detected by immunohistochemical staining; JNK mRNA levels were detected by PCR. Results:Compared with the model group, the serum levels of 25(OH)D3 and Omentin-1 in the Zhuanggu Zhitong Capsules group and the positive drug group increased ( P<0.01), the mean gray values of JNK and p-JNK protein in bone and kidney tissues decreased ( P<0.01), and the mRNA levels of JNK in bone and kidney tissues decreased ( P<0.01). Conclusion:Zhuanggu Zhitong Capsules can effectively improve the bone microstructure of postmenopausal osteoporotic rats, and its mechanism may be related to the regulation of JNK signaling pathway.
5.Clinical characteristics of juvenile dermatomyositis in anti-nuclear matrix protein 2 antibody-positive patients and risk factors for severity: a national multicenter retrospective study
Huiyuan YANG ; Wanzhen GUAN ; Ling2 YANG ; Haimei LIU ; Xiaoqing3 LI ; Haiguo YU ; Meiping LU ; Jun YANG ; Xiaohui LIU ; Hongxia ZHANG ; Wei ZHANG ; Jihong XIAO ; Xiaozhong LI ; Guomin LI ; Hong CHANG ; Sheng HAO ; Yue DU ; Daliang XU ; Ling WU ; Wenjie ZHENG ; Li LIU ; Xinhui JIANG ; Shaohui ZHU ; Dongmei ZHAO ; Xuemei TANG ; Li SUN
Chinese Journal of Pediatrics 2025;63(12):1299-1305
Objective:To investigate the clinical characteristics and independent risk factors of severe disease in patients with anti-nuclear matrix protein (NXP) 2 antibody-positive juvenile dermatomyositis (JDM).Methods:A retrospective cohort study was conducted, including 219 anti-NXP2 antibody-positive JDM patients admitted to 23 children′s hospitals across China from July 2011 to July 2023. Patients were classified into severe and non-severe groups based on classification criteria for severe dermatomyositis. Demographic characteristics, clinical manifestations, and laboratory parameters were compared between the 2 groups using independent sample t-test, Mann-Whitney U test, or χ2 test. Univariate and multivariate Logistic regression analyses were performed to identify risk factors for severe disease. The receiver operating characteristic curve was employed to calculate optimal cut-off values. Results:Among the 219 patients, 108 were male and 111 were female, with an age at onset of 6.3 (3.5, 9.4) years. The severe group comprised 69 patients, and the non-severe group 150 patients. The severe group had significantly higher rates of fever, heliotrope rash, subcutaneous edema, periorbital edema, anti-Ro52 antibody positivity, as well as elevated levels of ferritin-to-albumin ratio (FAR), creatine kinase (CK), aspartate aminotransferase (AST), and lactate dehydrogenase (LDH) (all P<0.05). Multivariate analysis identified anti-Ro52 antibody positivity ( OR=13.26, 95% CI 1.37-128.29) and elevated FAR ( OR=1.90, 95% CI 1.09-2.31) as independent risk factors for severe anti-NXP2 antibody-positive JDM (both P<0.05). Receiver operating characteristic curve analysis revealed that a FAR cutoff value of 6.82 predicted severe disease with an area under the curve of 0.87 (95% CI 0.81-0.94, P<0.001), sensitivity of 0.85, and specificity of 0.70. All patients received glucocorticoid therapy, and the severe group received higher proportions of steroid pulse therapy, cyclophosphamide, mycophenolate mofetil, intravenous immunoglobulin, biologics, and adjuvant treatments compared to the non-severe group (all P<0.05). In terms of outcomes, 2 patients (2.9%) in the severe group died (due to neurological involvement and intestinal perforation, respectively), while the remaining patients achieved complete clinical response or remission. All patients in the non-severe group achieved remission. Conclusions:The primary clinical features of anti-NXP2 antibody-positive JDM included fever, heliotrope rash, subcutaneous edema, periorbital edema, anti-Ro52 antibody positivity, and elevated levels of CK, AST, LDH, and FAR. Furthermore, anti-Ro52 antibody positivity and a FAR>6.82 were identified as independent risk factors.
6.Observation of the clinical efficacy of thermosensitive moxibustion on headache and dizziness in patients with sympathetic type of cervical spondylosis
Guomin HUANG ; Qiangjian MAO ; Lin YANG ; Ziru LI ; Yanan YANG ; Desheng WU ; Shuisheng ZHOU ; Zhenhai CHI
China Modern Doctor 2025;63(4):24-27
Objective To explore the clinical efficacy of thermosensitive moxibustion in treating headache and dizziness in patients with sympathetic type of cervical spondylosis(SCS).Methods A total of 80 patients with SCS treated in Affiliated Hospital of Jiangxi University of Chinese Medicine from June 2023 to June 2024 were selected and divided into observation group and control group according to random number table method,with 40 patients in each group.The patients in control group were treated with regular moxibustion,while the patients in observation group were treated with thermosensitive moxibustion.The clinical efficacy,pain,quality of life,cervical spine function,cervical sympathetic symptoms,and adverse reactions between two groups were compared.Results The total effective rate of observation group was significantly higher than that of control group(x2=4.501,P=0.034).After treatment,visual analogue scale(VAS),Northwick Park neck pain questionnaire(NPQ)and cervical sympathetic nerve symptom scores of patients in two groups were significantly lower than before treatment,and clinical assessment scale for cervical spondylosis(CASCS)scores were significantly higher than before treatment(P<0.05).VAS,NPQ and cervical sympathetic nerve symptom scores of observation group were significantly lower than those of control group,and CASCS score was significantly higher than that of control group(P<0.05).No adverse reactions such as fainting and scalding occurred in both groups during treatment.Conclusion Thermosensitive moxibustion for SCS can reduce cervical sympathetic nerve symptoms and pain,improve cervical spine function,and improve patients'quality of life,and is relatively safe.
7.Observation of the clinical efficacy of thermosensitive moxibustion on headache and dizziness in patients with sympathetic type of cervical spondylosis
Guomin HUANG ; Qiangjian MAO ; Lin YANG ; Ziru LI ; Yanan YANG ; Desheng WU ; Shuisheng ZHOU ; Zhenhai CHI
China Modern Doctor 2025;63(4):24-27
Objective To explore the clinical efficacy of thermosensitive moxibustion in treating headache and dizziness in patients with sympathetic type of cervical spondylosis(SCS).Methods A total of 80 patients with SCS treated in Affiliated Hospital of Jiangxi University of Chinese Medicine from June 2023 to June 2024 were selected and divided into observation group and control group according to random number table method,with 40 patients in each group.The patients in control group were treated with regular moxibustion,while the patients in observation group were treated with thermosensitive moxibustion.The clinical efficacy,pain,quality of life,cervical spine function,cervical sympathetic symptoms,and adverse reactions between two groups were compared.Results The total effective rate of observation group was significantly higher than that of control group(x2=4.501,P=0.034).After treatment,visual analogue scale(VAS),Northwick Park neck pain questionnaire(NPQ)and cervical sympathetic nerve symptom scores of patients in two groups were significantly lower than before treatment,and clinical assessment scale for cervical spondylosis(CASCS)scores were significantly higher than before treatment(P<0.05).VAS,NPQ and cervical sympathetic nerve symptom scores of observation group were significantly lower than those of control group,and CASCS score was significantly higher than that of control group(P<0.05).No adverse reactions such as fainting and scalding occurred in both groups during treatment.Conclusion Thermosensitive moxibustion for SCS can reduce cervical sympathetic nerve symptoms and pain,improve cervical spine function,and improve patients'quality of life,and is relatively safe.
8.Anti-3-hydroxy-3-methylglutaryl-coenzyme A reductase antibody-mediated necrotizing myopathy in 10 children: clinical features and treatment outcomes
Qing WU ; Qianying LYU ; Haimei LIU ; Wanzhen GUAN ; Yinyu GONG ; Yifan LI ; Qiaoqian ZENG ; Xiaomei ZHANG ; Qijiao WEI ; Ling YANG ; Guomin LI ; Xihua LI ; Lei ZHAO ; Li SUN
Chinese Journal of Rheumatology 2025;29(7):575-582
Objective:To analyze the clinical characteristics and treatment outcomes of children with anti-3-hydroxy-3-methylglutaryl-coenzyme A reductase (anti-HMGCR) antibody-mediated necrotizing myopathy, and to explore early identification and management strategies to provide reference for clinical diagnosis and treatment.Methods:A retrospective analysis was conducted on the clinical data and treatment outcomes of 10 pediatric patients with anti-HMGCR antibody-mediated necrotizing myopathy admitted to the Department of Rheumatology, Children′s Hospital of Fudan University from December 2020 to December 2024. Statistical description was performed using SPSS 22.0.Results:Among the 10 patients, the male-to-female ratio was 1:4, the age of onset was (7.2±4.0) years, and the disease duration at diagnosis was (22.2±19.6) months. None had a history of statin exposure. Six patients presented with muscle weakness, and4 were diagnosed due to asymptomatic elevation of creatine kinase (CK); 4 had dermatomyositis-like rashes. All patients showed significantly elevated CK levels [median 3 291(1 969, 8 776)U/L] and underwent muscle biopsy. Histopathological findings revealed myofiber degeneration, necrosis, and regeneration in all cases, with inflammatory infiltration in 9 cases, MHC-Ⅰ positivity in all, and C5b-9 positivity in 9 cases. The median follow-up duration was (15.7±6.3) months. At the last follow-up, muscle strength was normal or nearly normal, and the CK median value had decreased to 977.5 (211.0, 3 536.0) U/L.Conclusion:For patients with suspected idiopathic inflammatory myopathy and significantly elevated CK, muscle-specific antibody testing-including anti-HMGCR-and muscle biopsy should be performed promptly regardless of the presence of skin rash, to ensure accurate diagnosis and guide treatment, thereby avoiding misdiagnosis or missed diagnosis.
9.Repairment effect of intra-articular adipose stem cell injection on articular cartilage destruction in rabbit model with temporomandibular joint osteoarthritis and its mechanism
Jing HE ; Gao SUN ; Nannan LI ; Palizi ABULIKEMU ; Guomin WU
Journal of Jilin University(Medicine Edition) 2024;50(1):71-78
Objective:To discuss the repairment effect of intra-articular injection of adipose derived stem cells(ADSCs)on articular cartilage destruction in the temporomandibular joint osteoarthritis(TMJOA)model rabbits,and to clarify the possible mechanism.Methods:Twenty-seven rabbits were randomly divided into control group,model group,and ADSCs group.The ADSCs of the rabbits were extracted and cultured.The rabbit TMJOA model was prepared by monosodium-iodoacetate(MIA)injection technique.The temporomandibular joint cavity of the TMJOA model rabbits in ADSCs group was given two continuous intra-articular injections of 1.0×106 mL-1 ADSCs,while the rabbits in control and model group were given sequivalent volume of saline into the temporomandibular joint cavity.After 8 weeks,Micro-CT scan was performed on the temporomandibular joints of the rabbits in various groups;the bone volume fraction(BV/TV),bone surface area/bone volume(BS/BV),trabecular thickness(Tb.Th),trabecular separation(Tb.Sp),and trabecular number(Tb.N)of condyles tissue of the rabbits in various groups were analyzed;HE staining was used to observe the pathomorphology of condyles tissue of the rabbits in various groups;immunohistochemistry was used to detect the localization and expression levels of SRY-related high mobility group box gene 9(SOX9),matrix metalloproteinase-13(MMP-13),and vascular endothelial growth factor(VEGF)proteins in condyles tissue of the rabbits in various groups;Western blotting method was used to detect the expression levels of SOX9,MMP-13,and VEGF proteins in condyles tissue of the rabbits in various groups.Results:The micro-CT scan results showed that compared with control group,the BV/TV,Tb.Th,and Tb.N of condyles tissue of the rabbits in model group were significantly decreased(P<0.05),while the BS/BV and Tb.Sp were significantly increased(P<0.05);compared with model group,the BV/TV,Tb.Th,and Tb.N in condyles tissue of the rabbits in ADSCs group were significantly increased(P<0.05),and the BS/BV and Tb.Sp were significantly decreased(P<0.05).The HE staining results showed that the condylar cartilage surface of the rabbits in control group was smooth with clear layers and intact structure;compared with control group,the surface of condyles tissue of the rabbits in model group was irregular with thickened hypertrophic layer and areas of cell depletion and clustering;compared with model group,the pathological damage of condyles tissue of the rabbits in ADSCs group was significantly decreased.The immunohistochemical staining results showed that compared with control group and ADSCs group,the number of brown granule in condyles tissue of the rabbits in model group was increased,mainly concentrated in the hypertrophic layer,especially in the bone cartilage junction site and the expression levels of SOX9,MMP-13,and VEGF proteins in condyles tissue of the rabbits in model group were significantly increased(P<0.05);compared with model group,the number of brown granule in condyles tissue of the rabbits in ADSCs group was significantly decreased,and the expression levels of SOX9,MMP-13,and VEGF proteins were significantly decreased(P<0.05).The Western blotting results showed that compared with control group,the expression levels of SOX9,MMP-13,and VEGF proteins in condyles tissue of the rabbits in model group were significantly increased(P<0.05);compared with model group,the expression levels of SOX9,MMP-13,and VEGF proteins in condyles tissue of the rabbits in ADSCs group were significantly decreased(P<0.05).Conclusion:Intra-articular injection of ADSCs can effectively repair the cartilage destruction in TMJOA,alleviate the cartilage injury,and mitigate the progression of osteoarthritis.
10.Biological properties of concentrated growth factor combined with bone marrow mesenchymal stem cell sheet and its effect on bone defect repairment
Jianhong SHI ; Yuanye TIAN ; Kai CHEN ; Gao SUN ; Guomin WU
Journal of Jilin University(Medicine Edition) 2024;50(6):1535-1546
Objective:To discuss the effect of concentrated growth factor(CGF)on the performance of bone marrow mesenchymal stem cells(BMSCs)sheets,and to clarify the role of CGF-containing composite cell sheets(CS)in the bone defect repairment.Methods:In in vitro experiments,the BMSCs were isolated and cultured from two 3-week-old SD rats;Alizarin Red S and Oil Red O staining were used to identify the osteogenic and adipogenic capabilities of BMSCs;CGF liquid extracts(CGFe)was prepared from three 3-week-old SD rats.The cells were divided into control group,traditional CS(BMSC-CS)group,and CGF-containing composite CS(CGF/BMSC-CS)group.The morphology of the CS in two groups was observed by HE staining.Alizarin Red and alkaline phosphatase(ALP)staining were used to detect the osteogenic differentiation of the CS in various groups;cell scratch assay was used to detect the migration abilities of the cells in various groups;real-time fluorescence quantitative PCR(RT-qPCR)method was used to detect the mRNA expression levels of ALP,collagen are type 1(COL-1),Runt-related transcription factor 2(RUNX2),and osteocalcin(OCN)in the cells in various groups.In in vivo experiments,15 SD rats were randomly divided into control group,BMSC-CS group,and CGF/BMSC-CS group;micro computed tomography(Micro-CT)was used to detect the bone formation parameters in skull defects of the rats in various groups;HE staining and Masson staining were used to observe the morphology of skull defect tissue of the rats in various groups.Results:The third-generation BMSCs were spindle-shaped,closely arranged,and grew in a vortex cluster.The Alizarin red staining results showed obvious calcium nodules,and the Oil red O staining showed red lipid droplets,confirming the cells'ability to undergo osteogenic and adipogenic differentiation.The CS were white and semi-transparent,with slightly curled edges.The peeled CS were irregularly curled and wrinkled.Compared with BMSC-CS group,the CS in CGF/BMSC-CS group were whiter,less transparent,significantly increased in thickness and extensibility,less prone to breakage,and had a certain degree of stickiness and plasticity.The HE staining results showed that compared with BMSC-CS group,the number of the cells of CS in CGF/BMSC-CS group was increased,with denser arrangement and more abundant extracellular matrix(ECM),which wrapped and connected the cells to form an integral sheet-like structure.The Alizarin red and ALP staining results showed that compared with control group,the ALP activity and mineralization uplift value of CS in BMSC-CS group were significantly increased(P<0.05);compared with control group and BMSC-CS group,the number of osteoblasts and red mineralized nodules in the CS in CGF/BMSC-CS group was significantly increased,with obvious deepening of the staining,increased positive area,and the ALP activity and mineralization uplift value were significantly increased(P<0.05).Compared with BMSC-CS group,the ALP activity and mineralization uplif value of the CS in CGF/DMSC-CS group were increased(P<0.05).The cell scratch assay results showed that after 24 of culture,compared with control group,the migration rates of the cells in BMSC-CS group and CGF/BMSC-CS group were significantly increased(P<0.05).Compared with BMSC-CS group,the migration rate of the cells in CGF/BMSC-CS group was significantly increased(P<0.01).After 48 h of culture,compared with control group,the migration rate of the cells in CGF/BMSC-CS group was significantly increased(P<0.05).The RT-qPCR results showed that compared with control group,the expression levels of COL-1 and OCN mRNA in the cells in BMSC-CS group were significantly increased(P<0.01),and the expression levels of ALP,COL-1,OCN,and RUNX2 mRNA in the cells in CGF/BMSC-CS group were significantly increased(P<0.01).Compared with BMSC-CS group,the expression levels of ALP,COL-1,and OCN mRNA in the cells in CGF/BMSC-CS group were significantly increased(P<0.01).The Micro-CT detection results showed that in control group,the boundary of the rat skull defect area was clear,with almost no new bone formation.In BMSC-CS group,a small amount of new bone formed only at the edge of the bone defect in skull of the rats,with a significant gap in the central area of the defect.In CGF/BMSC-CS group,new bone formed along the edge of the bone defect towards the central area in skull of the rats,repairing most of the bone defect.Compared with control group,the bone volume(BV)and trabecular number(Tb.N)of the rats in BMSC-CS group were significantly increased(P<0.05);the bone volume(BV),bone volume fraction[BV/tissue volume(TV)],trabecular thickness(Tb.Th),and trabecular number(Tb.N)in skull of the rats in CGF/BMSC-CS group,were significantly increased(P<0.05).Compared with BMSC-CS group,the BV,BV/TV,Tb.Th,and Tb.N in skull of the rats in CGF/BMSC-CS group were significantly increased(P<0.01).The HE and Masson staining observation showed that in control group,almost no new bone formed in the skull defect tissue of the rats,with only a large amount of collagen fibers connecting the two sides of the bone ends.In BMSC-CS group,a small amount of new bone formed only at the edge of the bone defect in skull tissue of the rats,with the central area of the defect containing dense collagen fibers connected to the newly formed bone at the defect edge.In CGF/BMSC-CS group,new bone tissue could be seen at the edge of the bone defect,and bone islands formed in the central area of the defect,surrounded by osteocytes and a large amount of collagen fibers.The Masson staining observation results showed that the cytoplasm and osteoid were red,and the collagen was blue.In CGF/BMSC-CS group,newly formed osteoid was observed in skull defect tissue of the rats,with the highest amount of new bone formation.Conclusion:CGF can promote the osteogenic differentiation and increase the richness of ECM in BMSCs sheets.CGF-containing composite CS can efficiently repair skull defects of the rats and serve as an ideal and safe material for promoting the bone regeneration.

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