Objective To construct tissue-engineered skin via in vitro inoculation of epidermal stem cells(ESCS)onto de-epidermized dermis.Methods Skin tissue was obtained from the foreskin of a healthy 6-year-old child.and keratinocytes were isolated by two-step trypsinization method followed by the collection of ESCS via rapid adhesion by collagen Ⅳ.The ESCS were identified by morphological observation and immunohistochemical staining with K19 and integrin β1.To construct tissue-engineered skin,selected ESCS were seeded onto the surface of de-epidermized dermis followed by a one-week culture immersed in the medium and a subsequent 4-week culture at the air-medium interface.The tissue-engqneered skin was evaluated with haematoxylin & eosin(HE)staining as well as keratin immunohistochemistry.Results Micro scopically,cultured ESCs showed a paving stone-like appearance and grew into colonies.Immunohistochemistry revealed that the ESCs were positive for integrin-β1 and keratin 19.After 5 weeks of culture,3-6 layers of epidermal cell were observed on the dermis with the formation of stratum corneum.Keratin protein was observed in the artificial epidermal skin.Conclusion Tissue-engineered skin is successfully constructed with epidermal stem cells and de-epidermized dermis in vitro.

Objective To explore the effect of erythropoietin (EPO) attenuating apoptosis in old rat hippocampal neuronal cells exposed to sevoflurane and the role of toll like receptor 4.Methods Twenty months old SD rats,male,550-750 g,in accordance with the random number table,were divided into 3 groups (n =9):control group (group C),sevoflurane treatment (group S),and sevoflurane plus EPO treatment (group ES).The rats in group S and ES were subjected to inhale 4％ sevoflurane for 6 h,but the rats in group C were inhaled air-oxygen only.The rats in group ES were injected with EPO into caudal vein at 24 h,48 h,and 72 h after sevoflurane exposure.The cognitive ability was assessed by Morris water maze test;the effects of hippocampal apoptosis were assessed by TUNEL assays;the expressions of TLR4 mRNA was measured by RT-PCR assay;mitochondrial membrane potential (MMP) was assessed by JC-1 fluorescence;the expressions of APP and Aβ were assessed by western blot.Results Compared with group C,there were significant increases of escape latency period,neuronal apoptosis,TLR4 mRNA,and APP and Aβ expression,but a decrease of MMP in group S (P＜0.05).Compared with group S,there were significant decreases of escape latency period,neuronal apoptosis,TLR4 mRNA,and APP and Aβ expression,but a increase of MMP in group ES (P＜0.05).Conclusion The attenuation of rat hippocampal neuronal apoptosis induced by EPO could be associated with inhibition of TLR4,improvement of MMP,as well as inhibition of APP and Aβ activity.

Objective To evaluate the role of mitochondrial ATP sensitive potassium ( mito-KATP ) channel in dexmedetomidine-induced inhibition of subarachnoid hemorrhage ( SAH )-caused programmed cell death ( PCD) in cardiomyocytes of rats. Methods On hundred and twenty clean-grade healthy male Sprague-Dawley rats, aged 9-10 weeks, weighing 350-400 g, were divided into 5 groups ( n=24 each) using a random number table method: sham operation group ( group Sham ) , SAH group, SAH plus dexmedetomidine group ( group SD) , 5-HD plus SAH and dexmedetomidine group ( group HSD) and 5-HD plus SAH group ( group HS) . The rats were subjected to SAH by intracranial vascular puncture after being anesthetized with pentobarbital sodium. Dexmedetomidine 5 μg∕kg was infused for 10 min via the jugular vein starting from the time point after intracranial vascular puncture, followed by a continuous infusion of 5μg·kg-1 ·h-1 for 1 h in SD and HSD groups. 5-HD 30 mg∕kg was intraperitoneally injected at 1 h before intracranial vascular puncture in HSD and HS groups. Blood samples were collected from the abdominal aor-ta at 24 h after intracranial vascular puncture for determination of serum cardiac troponin I ( cTnI) concen-trations. The animals were then sacrificed, and myocardial specimens were collected for determination of PCD rate ( by TUNEL) , reactive oxygen species ( ROS) activity ( by DCFH-DA assay) , and expression of cleaved caspase-3, cleaved caspase-1 and interleukin-1beta ( IL-1β) ( by Western blot) . Results Com-pared with group Sham, the serum concentrations of cTnI, PCD rate and ROS activity were significantly in-creased, and the expression of cleaved caspase-3, cleaved caspase-1 and IL-1βwas up-regulated in SAH, SD, HSD and HS groups ( P<0. 05) . Compared with group SAH, the serum concentrations of cTnI, PCD rate and ROS activity were significantly decreased, and the expression of cleaved caspase-3, cleaved caspase-1 and IL-1βwas down-regulated in group SD, and the serum concentrations of cTnI, PCD rate and ROS activity were significantly increased, and the expression of cleaved caspase-3, cleaved caspase-1 and IL-1βwas up-regulated in group HS ( P<0. 05) . Compared with group SD, the serum concentrations of cT-nI, PCD rate and ROS activity were significantly increased, and the expression of cleaved caspase-3, cleaved caspase-1 and IL-1β was up-regulated in group HSD ( P<0. 05) . Compared with group HSD, the serum concentrations of cTnI, PCD rate and ROS activity were significantly increased, and the expression of cleaved caspase-3, cleaved caspase-1 and IL-1βwas up-regulated in group HS ( P<0. 05) . Conclusion mito-KATP channel is involved in dexmedetomidine-induced inhibition of PCD in cardiomyocytes of rats with SAH.

Objective To evaluate the role of Toll-like receptor 4 (TLR4) signaling pathway in sevoflurane anesthesia-induced cognitive dysfunction in aged rats.Methods Twenty-seven SPF healthy male Sprague-Dawley rats,aged 20 months,weighing 550-750 g,were divided into 3 groups (n =9 each) using a random number table:control group (C group),sevoflurane anesthesia group (S group) and TLR4 antagonist plus sevoflurane anesthesia group (TS group).TLR4 monoclonal antibody 30 μl was injected into the lateral cerebral ventricle in group TS,and the equal volume of serum containing no antibody was injected into the lateral cerebral ventricle in C and S groups.At 10 min after completion of injection,S and TS groups inhaled the mixture of 4％ sevoflurane and 30％ oxygen for 6 h,and group C only inhaled the mixture of air and oxygen.Morris water maze test was performed at 24 h after the end of sevoflurane anesthesia.The animals were sacrificed after completion of Morris water maze test,brains were removed and hippocampi were isolated for determination of nerve cell apoptosis (using TUNEL) and expression of activated caspase-3 (using immunofluorescent staining).Nerve cell apoptosis rate was calculated.The expression of high-mobility group box 1 protein (HMGB1) mRNA in hippocampi was measured by Northern blot assay at 6 h after the end of sevoflurane anesthesia.The expression of amyloid precursor protein (APP) and amyloid beta protein (Aβ) in hippocampi was assessed by Western blot at 24 h after the end of sevoflurane anesthesia.Results Compared with C group,the escape latency was significantly prolonged,nerve cell apoptosis rate was increased,the expression of activated caspase-3,HMGB1 mRNA,APP and Aβ was up-regulated in group S,and nerve cell apoptosis rate was increased,the expression of activated caspase-3,HMGB1 mRNA,APP and Aβ was up-regulated (P＜0.05),and no significant change was found in the escape latency in group TS (P＞0.05).Compared with S group,the escape latency was significantly shortened,nerve cell apoptosis rate was decreased,and the expression of activated caspase-3,HMGB1 mRNA,APP and Aβ was down-regulated in group TS (P＜0.05).Conclusion Activation of TLR4 signaling pathway is involved in the mechanism of sevoflurane anesthesia-induced cognitive dysfunction in aged rats.

Objective:To evaluate the application effect of vascularized iliac crest-tensor fascia lata flap(VIC-TFLF) based on deep circumflex iliac artery and vein in mandibular and oral soft tissue defect reconstruction.Methods:Retrospective analysis of the clinical data of patients with mandible and oral soft tissue defects in Henan Provincial People’s Hospital from October 2020 to March 2022. All cases were performed with computer-aided design and three-dimensional printing to make the models and guide plates. VIC-TFLF was used to repair and reconstruct the oral and mandible defects, the tensor fascia lata flap was used to repair the intraoral soft tissue defect, and the fascia lata was exposed to the mouth directly. After operation, the color, texture and change of intraoral flap were observed, and the recovery and complications were followed up.Results:In this study, 7 patients were included, 4 males and 3 females, aged 27-64 years old, with an average of 50.1 years old. There were 5 cases of squamous cell carcinoma of mandible gingival and cheek, 1 case of postoperative defect of pleomorphic adenocarcinoma of mandible, and 1 case of postoperative defect of ameloblastoma of mandible. According to the defect range of soft and hard tissue after resection, the flap areas of tensor fascia lata were 6.0 cm×3.0 cm-8.0 cm×6.0 cm, and the iliac bones were 3.7 cm×2.4 cm-9.2 cm×2.5 cm. All the composite tissue flaps survived, without distal necrosis, delayed wound healing and marginal fistula. The patients were followed up for 4-19 months, with an average of 11.7 months. The morphology and functions of the mandible and oral soft tissue were well recovered. The surface of tensor fascia lata was exposed to the oral cavity directly, which showed signs of mucosalization within 1 week after operation. The mucosalization was basically completed within 1 month, which was close to normal oral mucosal appearance, and could be reconstructed to produce better oral mucosal appearance in the later period. The wounds in the donor site healed well, and there was no abnormality in lower limb movement and thigh flexion. Among them, 3 patients had numbness on the lateral thigh skin of the donor site 3-5 days after operation. After 6 months of follow-up, the numbness in 2 cases disappeared, and 1 case was significantly reduced.Conclusions:The VIC-TFLF could obtain great appearance and function, with fewer complications and relatively small damage to the donor site, in repair and reconstruction of mandibular defects combined with oral soft tissue defects.

Objective:To evaluate the application effect of vascularized iliac crest-tensor fascia lata flap(VIC-TFLF) based on deep circumflex iliac artery and vein in mandibular and oral soft tissue defect reconstruction.Methods:Retrospective analysis of the clinical data of patients with mandible and oral soft tissue defects in Henan Provincial People’s Hospital from October 2020 to March 2022. All cases were performed with computer-aided design and three-dimensional printing to make the models and guide plates. VIC-TFLF was used to repair and reconstruct the oral and mandible defects, the tensor fascia lata flap was used to repair the intraoral soft tissue defect, and the fascia lata was exposed to the mouth directly. After operation, the color, texture and change of intraoral flap were observed, and the recovery and complications were followed up.Results:In this study, 7 patients were included, 4 males and 3 females, aged 27-64 years old, with an average of 50.1 years old. There were 5 cases of squamous cell carcinoma of mandible gingival and cheek, 1 case of postoperative defect of pleomorphic adenocarcinoma of mandible, and 1 case of postoperative defect of ameloblastoma of mandible. According to the defect range of soft and hard tissue after resection, the flap areas of tensor fascia lata were 6.0 cm×3.0 cm-8.0 cm×6.0 cm, and the iliac bones were 3.7 cm×2.4 cm-9.2 cm×2.5 cm. All the composite tissue flaps survived, without distal necrosis, delayed wound healing and marginal fistula. The patients were followed up for 4-19 months, with an average of 11.7 months. The morphology and functions of the mandible and oral soft tissue were well recovered. The surface of tensor fascia lata was exposed to the oral cavity directly, which showed signs of mucosalization within 1 week after operation. The mucosalization was basically completed within 1 month, which was close to normal oral mucosal appearance, and could be reconstructed to produce better oral mucosal appearance in the later period. The wounds in the donor site healed well, and there was no abnormality in lower limb movement and thigh flexion. Among them, 3 patients had numbness on the lateral thigh skin of the donor site 3-5 days after operation. After 6 months of follow-up, the numbness in 2 cases disappeared, and 1 case was significantly reduced.Conclusions:The VIC-TFLF could obtain great appearance and function, with fewer complications and relatively small damage to the donor site, in repair and reconstruction of mandibular defects combined with oral soft tissue defects.

Objective:To introduce the clinical application of three-dimensional(3D) printed titanium alloy prosthesis in the reconstruction of mandibular defects.Methods:Three patients with extensive mandibular defects, who were either ineligible for or refused autologous bone transplantation were selected from the Oral and Maxillofacial-Head and Neck Oncology Department at School & Hospital of Stomatology, Wuhan University between April 2019 and December 2021. Preoperative oral and maxillofacial CT scanning was performed, followed by a virtual surgical plan and the design and manufacture of titanium mandibular prosthesis as well as the surgery guide-plate. Repair of mandibular defects using 3D printed titanium mandibular prosthesis. Follow-up after surgery to evaluate the patient’s functional and cosmetic recovery.Results:The study included three male patients aged 27, 10 years and 8 months, and 74 years, respectively. Two cases involved recurrent mandibular tumors postoperatively, and one case involved mandibular defects following gingival tumor surgery that affected the patient’s facial appearance and eating. All surgeries were successfully completed, with postoperative outpatient follow-ups at 36, 32, and 6 months, respectively. Follow-up indicated that the facial contours were basically symmetrical, the degree of mouth opening was normal, and the occlusion on the healthy side was essentially normal. No exposure, loosening, or fracturing of the prostheses was observed.Conclusion:3D printed titanium mandibular prosthesis can effectively restore the facial contour and mandibular function of patients.

Objective:To introduce the clinical application of three-dimensional(3D) printed titanium alloy prosthesis in the reconstruction of mandibular defects.Methods:Three patients with extensive mandibular defects, who were either ineligible for or refused autologous bone transplantation were selected from the Oral and Maxillofacial-Head and Neck Oncology Department at School & Hospital of Stomatology, Wuhan University between April 2019 and December 2021. Preoperative oral and maxillofacial CT scanning was performed, followed by a virtual surgical plan and the design and manufacture of titanium mandibular prosthesis as well as the surgery guide-plate. Repair of mandibular defects using 3D printed titanium mandibular prosthesis. Follow-up after surgery to evaluate the patient’s functional and cosmetic recovery.Results:The study included three male patients aged 27, 10 years and 8 months, and 74 years, respectively. Two cases involved recurrent mandibular tumors postoperatively, and one case involved mandibular defects following gingival tumor surgery that affected the patient’s facial appearance and eating. All surgeries were successfully completed, with postoperative outpatient follow-ups at 36, 32, and 6 months, respectively. Follow-up indicated that the facial contours were basically symmetrical, the degree of mouth opening was normal, and the occlusion on the healthy side was essentially normal. No exposure, loosening, or fracturing of the prostheses was observed.Conclusion:3D printed titanium mandibular prosthesis can effectively restore the facial contour and mandibular function of patients.