Objective:To construct human adipose-derived mesenchymal stem cells (hASCs)-biomate-rial mixture 3D bio-printing body and detect its osteogenesis in vivo,and to establish a guideline of osteogenesis in vivo by use of 3D bio-printing technology preliminarily.Methods:P4 hASCs were used as seed cells,whose osteogenic potential in vitro was tested by alkaline phosphatase (ALP)staining and alizarin red staining after 1 4 d of osteogenic induction.The cells were added into 20 g/L sodium alginate and 80 g/L gelatin mixture (cell density was 1 ×1 06/mL),and the cell-sodium alginate-gelatin mixture was printed by Bioplotter 3D bio-printer (Envision company,Germany),in which the cells’survival rate was detected by live-dead cell double fluorescence staining.Next,the printing body was osteogeni-cally induced for 1 week to gain the experimental group;and the sodium alginate-gelatin mixture without cells was also printed to gain the control group.Both the experimental group and the control group were implanted into the back of the nude mice.After 6 weeks of implantation,the samples were collected,HE staining,Masson staining,immunohistochemical staining and Inveon Micro CT test were preformed to analyze their osteogenic capability.Results:The cells’survival rate was 89%±2% after printing.Six weeks after implantation,the samples of the control group were mostly degraded,whose shape was irregu-lar and gel-like;the samples of the experimental group kept their original size and their texture was tough.HE staining and Masson staining showed that the bone-like tissue and vessel in-growth could be observed in the experimental group 6 weeks after implantation,immunohistochemical staining showed that the result of osteocalcin was positive,and Micro CT results showed that samples of the experimental group had a higher density and the new bone volume was 1 8%±1%.Conclusion:hASCs-biomaterial mixture 3D bio-printing body has capability of ectopic bone formation in nude mice,and it is feasible to apply cells-biomaterial mixture 3D bio-printing technology in the area of bone formation in vivo.

Objective To investigate the outcome of free posterolateral leg perforator flap in the reconstruction of soft tissue defects of oral and maxillofacial area.Methods From February,2014 to August,2016,16 patients were performed defect reconstruction following oral cancer ablation with free posterolateral leg perforator flap including 3 cases of sequamous cell carcinoma of the tongue,6 cases of carcinoma of the buccal mucosa,7 cases of sequamouse cell carcinoma of the floor of mouth.The flap ranged from 5.0 cm×3.0 cm to 13.0 cm ×9.0 cm in size,and were adjected to the soft tissue defects of oral and maxillofacial area.The outcome of reconstruction was evaluated by considering the facial appearance,the swallowing and the speach function,the ranges of mouth opening and patient prognosis.Results All 15 transplanted flaps survived well and only 1 survived after disposing in time,because of vascular crisis.The donor sites were closed directly without donor-site morbidity.All patients were satisfactory with their facial appearance,the ranges of mouth opening and swallowing and speech function.Conclusion The free posterolateral leg perforator was an ideal free tissue for repairing functionally oral and maxillofacial defects,sinceit including some beneficial characteristic such as constant blood vessel,flexible and divese design,abundant and position-latent donor site tissues,small operation loss,flexible preparation and loss of major vascular nerves.

Objective To observe the clinical effects of unilateral cleft lip repairmen via improved Millard technique. Methods 64 cases of patients with unilateral cleft lip were collected from July 2014 to June 2015. The traditional Millard technique was improved by moving traditional fixed point of columella nasi to close with the lateral nasal columella roots,and the moving range was adjusted according to the nasal columella deformity and nostril size. Meanwhile ,the nasal floor and the alar basal incisions were not designed. Instead ,fully free of affected side orbicularis oris muscles were applied to making apposition suture with the healthy side. Results It showed that anatomic form of the upper lip and the philtral column of patients ,whose unilateral cleft lips were repaired by the improved Millard technique,were normal,and both sides of the lip peak were basically symmetrical after 1 ~ 12 month(s)follow-up. Conclusions Compared with the traditional Millard technique,the improved technique could bring a better effect on appearance of the patients. And it will be one of the ideal repairing techniques for unilateral cleft lip.

Previous studies have suggested that various kinds of inflammatory factors can influence the formation and development of tumor cells.Researche has shown that gallbladder cancer is closely linked with local inflammation,which is a risk factor for the development of gallbladder cancer.It is widely known that cholecystitis is closely correlated with gallstones,and that bile obtained from patients with gallbladder cancer contains a large variety of bacteria,such as Salmonella typhi,Helicobacter,and Escherichia coli.It is proposed that the gallbladder may be the result of the joint action of inflammation with the bacterial flora.Similarly,the inflammatory “tumor infiltrating lymphocyte” (TIL)can be observed in the tumor and its surrounding tissues,and may also play a role in tumor growth and metastasis.However,detailed mechanisms about the relationship between inflammation and gallbladder cancer is still not clear.No specific anti-inflammatory drugs for gallbladder cancer have been developed. In the near future,anti inflammatory drugs may play a more important role in gallbladder cancer prevention and treatment.

Objective:To investigate the effects of ultrasound-guided lumbar erector spinae plane block (L-ESPB) combined with general anesthesia using a laryngeal mask airway on postoperative delirium and early outcome in older adult patients undergoing hip arthroplasty.Methods:Sixty older adult patients who underwent unilateral total hip arthroplasty under general anesthesia using a laryngeal mask airway from August 2020 to May 2021 were included in this study. They were randomly assigned to observation and control groups, with 30 patients in each group. All patients underwent general anesthesia using a laryngeal mask airway. In the observation group, L-ESPB with 30 mL 0.375% ropivacaine was performed before induction of general anesthesia. Patient-controlled intravenous analgesia was performed in all patients after surgery. The Visual Analogue Scale score was > 4 points, and dezocine was intravenously injected for rescue analgesia. The amount of general anesthetics, the rate of use of vasoactive drugs, the time to pull out the laryngeal mask airway, length of stay in postanesthesia care unit, and the incidence of restlessness during the recovery period were recorded. Resting-state VAS score and Bruggrmann comfort scale score at 4, 8, 12, 24 and 48 hours after surgery, effective times of pressing the analgesia pump, the rate of rescue analgesia, the incidence of anesthesia-related adverse reactions at 48 hours after surgery, Pittsburgh Sleep Quality Index (PSQI) score at 1 day before and 1, 3 and 5 days after surgery, the incidence of postoperative delirium within 5 days after surgery, the time to get out of bed the first time, and the incidence of pulmonary infection were recorded.Results:The amount of remifentanil and propofol used during surgery were significantly lower in the observation group than those in the control group ( t = -6.80, -5.23, both P < 0.05). The rate of use of urapidil and esmolol were significantly lower in the observation group than those in the control group ( χ2 = 4.32, 5.46, both P < 0.05). The time to pull out the laryngeal mask airway, length of stay in postanesthesia care unit, and the time to get out of bed the first time in the observation group were (14 ± 2) minutes, (21 ± 2) minutes and (2.4 ± 0.5) days respectively, which were significantly shorter than those in the control group [(18 ± 2) minutes, (26 ± 3) minutes and (2.8 ± 0.7) days, t = -6.64, -7.18, -2.51, all P < 0.05]. The incidence of restlessness during the recovery period, postoperative delirium, and pulmonary infection in the observation group were 0.0%, 3.3% and 0.0% respectively, which were significantly lower than those in the control group [20.0%, 26.7%, 20.0%, all P < 0.05]. Resting-state VAS score at each time point was significantly lower in the observation group than that in the control group ( t = 3.32, 2.97, 4.33, 3.81, 3.10, all P < 0.05). Bruggrmann comfort scale score at each time point was significantly higher in the observation group than that in the control group ( t = 9.20, 8.62, 5.73, 5.72, 6.28, all P < 0.05). The effective times of pressing the analgesia pump, the rate of rescue analgesia, and the incidence of nausea and vomiting at 48 hours after surgery in the observation group were 0 (0, 0), 10.0% and 6.7% respectively, which were significantly lower than those in the control group [1(0, 2), 33.3% and 30.0%, Z = -3.41, χ2 = 4.81, 5.46, all P < 0.05]. PSQI score measured at 1, 3, and 5 days after surgery were significantly increased compared with those measured 1 day before surgery in each group ( F = 116.80, 325.64, both P < 0.001). PSQI measured at 1, 3 and 5 days after surgery was significantly lower than those in the control group ( t = -7.26, -6.41, -11.17, all P < 0.001). There were no significant differences in the incidence of dizziness, drowsiness, urinary retention and atelectasis between the two groups ( P > 0.05). Conclusion:Ultrasound-guided L-ESPB combined with general anesthesia using a laryngeal mask airway for hip arthroplasty in older adult patients can decrease the incidences of postoperative delirium and complications, which are conducive to early postoperative outcomes.

Objective Mouse models of sepsis-induced myocardial injury by intraperitoneal injection of lipopolysaccharide (LPS) was established in order to provide a reliable method for the research of pathogenesis of sepsis-induced myocardial injury. Methods According to the method of random number table, a total of 150 male C57BL/6 mice were divided into five groups: NC group, sham group, and LPS 10, 12, 15 mg/kg groups, with 30 in each group. Septic myocardial injury was induced by intraperitoneal injection LPS in mice; sham group was injected with equal 0.9% saline; while there was no treatment in mice of NC group. Fifteen of the 30 mice in each group were used to observe the general status of mice before and after LPS or saline injection. Twenty-four hours after LPS or saline injection, the left ventricular function was assessed by echocardiography, serum level of cardiac troponin (cTnI) was determined by enzyme linked immunosorbent assays (ELISA), and the cardiac histomorphology and ultrastructure were observed; the other 15 mice were used to monitor the 7-day mortality after LPS or saline injection. Results The mice challenged to LPS displayed symptoms of sepsis, such as depression, ruffled fur, and diarrhea. Compared with NC group, left ventricular ejection fraction (LVEF), left ventricular fraction shortening (LVFS) were significantly decreased at 24 hours after LPS administration in LPS 10, 12, 15 mg/kg groups [LVEF: 0.459±0.044, 0.432±0.034, 0.348±0.064 vs. 0.588±0.019, LVFS: (22.36±2.60)%, (20.78±1.91)%, (16.27±3.31)% vs. (30.55±1.30)%, all P < 0.01], and cTnI levels were significantly increased (ng/L: 270.40±43.50, 281.14±41.79, 298.39±42.05 vs. 192.59±16.90, all P <0.01). Myocardium injury was observed in three LPS groups, myocardial fibrosis, interstitial edema, erythrocyte leakage and infiltrating inflammatory cells were observed under light-microscope; ultrastructural changes disorderly arranged in cardiac muscle fibers, mitochondrial swelling and even partly missing mitochondria cristae were found under transmission electron microscope (TEM), and the higher of the dose, the more sever of the damage. There was no significant difference between sham group and NC group. The 7-day mortality in LPS 10, 12, 15 mg/kg groups were 33.3%, 53.3% and 86.7%, respectively, while no death in the NC group and sham group. Conclusion For establishing the mouse model of sepsis-induced myocardial injury, intraperitoneal injection with 12 mg/kg LPS is a preferable choice in our research.

Objective To evaluate the effect of double-leaf perforator free flap pedicled with posterolateral calf peroneal artery on repairing facial through-and-through defect after oral cancer oblation.Methods Nine patients with facial through-and-through defects after oblation of oral and maxillofacial malignancies,including 4 cases of buccal mucosa carcinoma,2 cases of maxilla osteosarcoma,1 case of carcinoma of the maxillary sinus and 2 cases of parotid duct carcinoma,were recruited from May,2016 to May,2018.The flap was used to repair the facial defect of recruited patients.The area of the bigger leaf of bilobed perforator flap was 7.0 cm×8.0 cm-6.0 cm×7.0 cm and the small leaf was 4.5 cm×4.5 cm-4.0 cm×4.0 cm,respectively.The intraoral and facial defects needed to be reconstructed contained the oral mucosal of the upper palate,the skin,subcutaneous tissue and mucosa of the cheek.The radiotherapy was performed 1.0-1.5 months after the operation.The prognosis including appearance,mouth opening,and the functions of deglutition,and language were assessed.Results Through 3-18 months outpatient followed-up,all 9 cases of transplanted flaps survived well and the incisions of the donor and recipient areas healed by first intention.The patients were satisfied with the facial appearance,mouth opening,and the functions of deglutition and language at the 6 months follow-up.The radiotherapy had no damage on the survival of the bilobed perforator flap.There was no recurrence or metastasis in follow-up.Conclusion The double-leaf perforator free flap pedicled with posterolateral calf peroneal artery is an ideal free tissue for repairing the facial perforating defect after oral cancer oblation because of its constant blood vessel,abundant tissues,flexible design and operating methods.

To evaluate the clinical effect of application of free peroneal artery perforator flap in repairing postoperative tissue defect after resection of oral maxillary malignant tumor. Methods From January, 2016 to June, 2018, 14 patients with maxillary malignant tumor were treated with tumor excision. Then the free per-oneal artery perforator flap was used to reconstruct tissue defect caused by tumorectomy. There were 6 cases of squa-mous-cell carcinoma of palatine, 2 cases of adenoid cystic carcinoma of palatine, and 6 cases of maxillagingiva squa-mous cell carcinoma. The incised area of flap was 4.0 cm×5.0 cm to 7.0 cm×8.0 cm. There were 5 cases of hard palate soft tissue defect repair and 9 cases of simultaneous repair of soft and hard palate. Followed-up by outpatient service in 3-12 months after operation, postoperative maxillamorphology, phonetic function, swallow function, opening degree and prognosis of the patients were evaluated. Results All 14 implanted flaps were alive.One flap had vas-cular crisis, and rescued by surgical exploration and timely rescue.Three flaps were prolonged healing.All donor sites were sutured directly.All surgical incisions healed primarily.Half a year after the operation, the appearance of maxilla was formed gradually. The phonetic function, swallowing function, opening degree and movement of lower leg were all recovered normal. One year after the operation, epithelization was done in 6 cases. And there was no tumor recur-rence.Conclusion The peroneal artery perforator flap has long vascular pedicle, larg diameter, high survival rate after vascular anastomosis and relatively concealed donor area. It can be used to repair and reconstruct the tissue defect in maxillary malignant tumor resection and achieved good result.

Objective To explore the effect of knocking down Rho-associated coiled-coil kinase (ROCK2) gene on the cognitive function of amyloid precursor protein/presenilin-1 (APP/PS1) double transgenic mice and its mechanism. Methods APP/PS1 double transgenic mice were randomly divided into AD model group (AD group), ROCK2 gene knock-down group (shROCK2 group), ROCK2 gene knock-down control group (shNCgroup), and wild-type C57BL/6 mice of the same age served as the wild-type control (WT group). Morris water maze and Y maze were employed to test the cognitive function of mice. Neuron morphology was detected by Nissl staining. Immunofluorescence histochemical staining was used to detect the expression of phosphorylated dynamin-related protein 1 (p-Drp1) and mitochondrial fusion 1 (Mfn1). Western blot analysis was used to detect the expression ROCK2, cleaved-caspase-3 (c-caspase-3), B-cell lymphoma 2 (Bcl2), Bcl2-related protein X (BAX), p-Drp1, mitochondrial fission 1 (Fis1), optic atrophy 1 (OPA1), Mfn1 and Mfn2. Results Compared with AD group mice, the expression of ROCK2 in shROCK2 group mice was significantly reduced; the cognitive function was significantly improved with the number of neurons in the hippocampal CA3 and DG areas increasing, and nissl bodies were deeply stained; the expression of c-caspase-3 and BAX was decreased, while the expression of Bcl2 was increased; the expression of mitochondrial division related proteins p-Drp1 and Fis1 were decreased, while the expression of mitochondrial fusion-related proteins OPA1, Mfn1 and Mfn2 were increased. Conclusion Knock-down of ROCK2 gene can significantly improve the cognitive function and inhibit the apoptosis of nerve cells of APP/PS1 mice. The mechanism may be related to promoting mitochondrial fusion and inhibiting its division.
Animals ; Mice ; Alzheimer Disease/pathology* ; Amyloid beta-Peptides/metabolism* ; Amyloid beta-Protein Precursor ; Apoptosis/genetics* ; bcl-2-Associated X Protein ; Caspase 3 ; Cognition ; Disease Models, Animal ; Mice, Inbred C57BL ; Mice, Transgenic ; Mitochondrial Dynamics/genetics*

【Objective】 To construct the Rb luciferase reporter gene assay system and detect the activation ability of Rb gene for screening the targeted drugs. 【Methods】 The synthetic Rb gene sequence was annealed to form a double-stranded DNA structure and then inserted into the polyclonal site of pGL6-TA. The junction product was transformed into E.coli DH5α competent cells for expanded culture, and the constructed pGL6-Rb-Luc plasmid and pGL6-TA plasmid were transfected into HEK293 cells. The monoclonal cell line HEK293-Rb-Luc with stable expression was screened by G418, and the activation and inhibition of Rb in HEK293-Rb-Luc were tested by serum and CDK4/6 inhibitor Palbociclib. 【Results】 The sequence of Rb reaction elements in pGL6-Rb-Luc was completely correct. The recovery of serum culture significantly increased the luciferase activity in HEK293-Rb-Luc (P<0.001). Compared with 0 nmol/L, 25, 50, 75 and 100 nmol/L, CDK4/6 inhibitor Palbociclib made the inhibition rate of Rb activity rise to 6.90%, 40.23%, 50.57% and 52.07%, respectively (P<0.05). 【Conclusion】 The Rb luciferase reporter gene detection system HEK293-Rb-Luc was successfully constructed, which can effectively detect the activation level of Rb.