Objective: To investigate the effect of icaritin on maturation and mineralization of mouse osteoblast MC3T3-E1 cells and its mechanism. Methods: The cultured MC3T3-E1 cells were divided into blank control group, CXC chemokine receptor type 4 (CXCR4) inhibitor (AMD3100) group, icaritin group, and icaritin plus AMD3100 group. The expression of CXCR4, stromal cell-derived factor 1 (SDF-1) and osteogenesis-related genes and proteins were detected by real-time RT-PCR and Western blotting after drug treatment for 24 h. The alkaline phosphatase (ALP) activity was determined with ALP kit on d3 and d6; calcium nodules were detected by alizarin red staining after drug treatment for 14 d. Results: Real time RT-PCR showed that compared with the blank control group, relative expressions of CXCR4, SDF-1 and osteogenesis-related genes in icaritin group were significantly increased (P<0.05 or P<0.01); After AMD3100 treatment, the relative expression of CXCR4 gene was decreased (P<0.05). Western blot showed that compared with the blank control group, relative expressions of CXCR4, SDF-1 and osteogenesis-related proteins in the icaritin group were significantly increased (all P<0.01), but were decreased after AMD3100 was added (all P<0.01). The ALP activity of icaritin group was significantly higher than that of blank control group (all P<0.01) on d3 and d6 after drug treatment, while the activity of ALP was significantly decreased after AMD3100 treatment (all P<0.01). At d14 after drug treatment, compared with the blank control group, the area of alizarin red staining was increased in the icaritin group, while it was significantly reduced after the addition of AMD3100. Conclusion: Icaritin may promote maturation and mineralization of mouse osteoblast MC3T3-E1 cells through CXCR4/SDF-1 signaling pathway.

Objective: To investigate the effect of resveratrol on peak bone mineral density and bone mass in growing rats. Methods: Thirty-six female healthy Wistar rats were randomly divided into control group, icariin group and resveratrol group with 12 rats in each group. Icariin (25 mg·kg^-1·d^-1), resveratrol (8.4 mg·kg^-1·d^-1) or equal volume of distilled water were given by gavage to icariin group, resveratrol group and control group, respectively. The rats were sacrificed after 12 weeks. The organ indexes were calculated and pathology sections were observed; the bone mineral density (BMD), bone biomechanics, serum bone metabolism index, and results of micro-CT scan were analyzed. Results: During the experiment, the body weight of rats showed an increasing trend and there was no significant difference among three groups (P>0.05). There were no significant differences in organ index of vital organs and pathological changes among the groups (all P>0.05). Compared with the control group, the whole body BMD, and the BMDs of femur and vertebrae in icariin and resveratrol groups were significantly increased after 12 weeks (all P<0.05). The maximum load values of femur and vertebrae, as well as elastic modulus of vertebrae in icariin and resveratrol groups were significantly higher than those in control group (P<0.05 or P<0.01). Micro-CT scan showed that the volumetric BMD, number of trabecular, trabecular thickness and bone volume/tissue volume of the cancellous bone in icariin and resveratrol groups were significantly higher and the trabecular separation was significantly lower than those in the control group (P<0.05 or P<0.01); while there was no significant difference in volumetric BMD of cortical bone for femur. The contents of osteocalcin in icariin and resveratrol groups were significantly higher than those in control group (all P<0.05), while the contents of tartarte-resistant acid phosphatase 5b (TRACP5b) were significantly lower than those in control group (all P<0.05).Conclusion: Resveratrol can inhibit bone resorption and enhance bone formation, so as to improve the peak bone mass and bone density, enhance bone strength and improve the microstructure of bone tissue in young rats.

Objective: To compare the effects of 50 Hz 1.8 mT sinusoidal magnetic field (SEMF) and 50 Hz 0.6 mT pulsed electromagnetic field(PEMF) on the maturation and mineralization of rat calvaria osteoblasts. Methods: Primary cultured rat calvarial osteoblasts were divided into 3 groups:blank control group, SEMF group and PEMF group. The rats in SEMT and PEMT groups were treated with 50 Hz 1.8 mT SEMF or 50 Hz 0.6 mT PEMF for 90 min/d, respectively. Western blotting and Real-time RT-PCR were used to detect the protein and mRNA expressions of Collagen-1, bone morphogenetic protein 2 (BMP-2), osterix (OSX) and Runt-associated transcription factor 2(Runx-2). The alkaline phosphatase(ALP) activity was detected by ALP test kits at d6 and d9 after treatment, and by ALP staining using azo coupling at d10 after treatment. The formation of calcium nodules was observed by alizarin red staining. Results: Compared with blank control group, the protein and mRNA expressions of Collagen-1, BMP-2, OSX and Runx-2 in SEMT and PEMT groups were significantly increased (P < 0.01 or P < 0.05); while the mRNA expressions of Collagen-1 and BMP-2 in PEMF group were significantly higher than those in SEMF group. After 6 days treatment, the activity of ALP in PEMF group was significantly higher than that in blank control group (P < 0.05), while such difference was not observed in SEMF group (P>0.05); after 9 days treatment, the activities of ALP in both PEMF and SEMP groups were significantly higher than that in blank control group (all P < 0.05), but the difference between PEMF and SEMF groups was not significant (P>0.05). After 10 days treatment, ALP staining was increased in both PEMF and SEMF groups compared with that in blank control group (all P < 0.01), and the stained area was bigger in PEMF group than that in SEMF group (P < 0.05). After 12 days treatment, calcium nodules were increased in PEMF and SEMF groups compared with that in blank control group (all P < 0.01), and more calcium nodules were observed in PEMF group than SEMF group (P < 0.05). Conclusion: Both 50 Hz 1.8 mT that in SEMF and 50 Hz 0.6 mT PEMF can promote the maturation and mineralization of osteoblasts, and the effect of PEMF is more marked.

Objective: To analysis the biomechanical and biocompatible properties of calcium phosphate cement (CPC) enhanced by chitosan short nanofibers(CSNF) and Arg-Gly-Asp (RGD). Methods: Chitosan nanofibers were prepared by electrospinning, and cut into short fibers by high speed dispersion. CPC with calcium phosphorus ratio of 1.5:1 was prepared by Biocement D method. The composition and structure of CPC, CSNF, RGD modified CSNF (CSNF-RGD), CSNF enhanced CPC (CPC-CSNF), RGD modified CPC-CSNF (CPC-CSNF-RGD) were observed by infrared spectrum, X-ray diffraction (XRD) and scan electron microscopy (SEM). The mechanical properties were measured by universal mechanical testing instrument. The adhesion and proliferation of MC3T3 cells were assessed using immunofluorescence staining and MTT method. Results: The distribution of CSNF in the scaffold was homogeneous, and the porous structure between the nanofibers was observed by SEM. The infrared spectrum showed the characteristic peaks at 1633 nm and 1585 nm, indicating that RGD was successfully grafted on chitosan nanofibers. The XRD pattern showed that the bone cement had a certain curability. The stain-stress test showed that break strengths were (17.74±0.54) MPa for CPC-CSNF and (16.67±0.56) MPa for CPCP-CSNF-RGD, both were higher than that of CPC(all P < 0.05). The immunofluorescence staining and MTT results: indicated that MC3T3 cells grew better on CPC-CSNF-RGD after 240 min of culture(all P < 0.05). Conclusion: CSNF-RGD can improve the biomechanical property and biocompatibility of CPC, indicating its potential application in bone tissue repair.

Objective: To evaluate the application of mechanically reinforced 45S5 Bioglass^®-derived glass ceramic porous scaffolds for repair of bone defect in rabbits. Methods: The BG-ZnB powders were added into the 45S5 Bioglass^® powder/paraffin microsphere mixtures and were sintered at 900℃ to obtain porous scaffolds with highly bioactive BG-ZnB of 0%, 2% or 4% of mass fraction (denoted as 45S5/ZnB0, 45S5/ZnB2, 45S5/ZnB4). Phase composition, porosity and compression properties of three kinds of as-sintered scaffolds were characterized by X-ray analysis, mercury porosimetry, and mechanical test. Thirty-six male New Zealand rabbits with critical-sized femoral bone defects were randomly divided into three groups (45S5/ZnB0 group, 45S5/ZnB2 group and 45S5/ZnB4 group, 12 for each), and were implanted with three kinds of porous scaffolds respectively. X-ray, micro-CT three-dimensional reconstruction and tissue slice staining were used to detected the efficiency of bone regeneration at 6 and 16 weeks after operation. The growth of newly formed bone was observed using HE, Masson staining and EnVision method. Results: Phase compositions of 45S5/ZnB2 and 45S5/ZnB4 were the same with 45S5/ZnB0, but the average pore size and porosity of the scaffolds were decreased with the increase of BG-ZnB content. 45S5/ZnB2 and 45S5/ZnB4 scaffolds exhibited higher compressive strength, osteogenesis and trabecular density than those of the 45S5/ZnB0 scaffold (all P < 0.05). With the mechanical reinforcement of BG-ZnB increased, the content of new bone, collagen type I and osteocalcin increased. Conclusion: Low-melt BG-ZnB-assisted sintering is a promising approach to improve the mechanical strength of 45S5 Bioglass^®.

Objective: To explore the effect of microRNA(miR)-222 on cell proliferation and apoptosis of fibroblasts in hypertrophic scar (HS) and the underlying mechanisms. Methods: The expression of miR-222 in the HS and the normal skin tissues was detected by real-time RT-PCR. The HS fibroblasts were transfected with miR-222 mimic and miR-222 inhibitor respectively. The cell viability was tested with MTT assay, cell cycle distribution and apoptosis were detected with flow cytometry and the expression levels of proliferation, apoptosis and cell cycle related proteins were determined with Western blot. Direct target of miR-222 was evaluated by dual-luciferase reporter assay. Results: miR-222 was significantly up-regulated in HS tissues compared with normal skin tissues(P < 0.05). Overexpression of miR-222 enhanced the cell viability of HS fibroblasts; increased mRNA and protein expressions of proliferating cell nuclear antigen (PCNA), collagen alpha-1 (Ⅰ) chain (Col1A1) and collagen alpha-1 (Ⅲ) chain (Col3A1); increased cell population in S phase and protein expressions of cyclin D1, cyclin E1 and cyclin-dependent kinases 1 (CDK1); inhibited cell apoptosis and reduced protein expressions of caspase-3/9. Overexpression of MMP1 attenuated the effects of miR-222 on the cell viability and apoptosis in fibroblasts, reduced expression levels of PCNA, cyclin D1 and the expression of caspase-3 was increased. Conclusion: miR-222 enhances cell proliferation and inhibits cell apoptosis of HS fibroblasts through negative regulation of MMP1, which suggests that miR-222 and MMP1 might be used as novel biomarkers and targets in diagnostic and therapeutic approaches for HS.

Objective: To investigate the function of primary cilium as an oxygen sensor in PC12 cells. Methods: The PC12 cells were transfected with IFT88 siRNA. The nuclear translocation of hypoxia inducible factor-1α (HIF-1α), nuclear factor erythroid-2 related factor 2 (Nrf2), and ciliogenesis were observed by immunofluorescence staining; and the mRNA expressions of HIF-1α, Nrf2, vascular endothelial growth factor (VEGF) and superoxide dismutase (SOD) were detected by real-time RT-PCR. Results: The ciliogenesis was inhibited in PC12 cells transfected with IFT88 siRNA. In hypoxia group and scramble control group, nuclear translocations of HIF-1α and Nrf2 were observed and mRNA expressions of HIF-1α, Nrf2, VEGF were increased, and those of SOD were decreased. While in PC12 cells transfected with IFT88 siRNA, nuclear translocations of HIF-1α and Nrf2 were not observed, and mRNA expressions of HIF-1α, Nrf2, VEGF were inhibited, and mRNA expression of SOD was increased. Conclusion: Primary cilia may act as an oxygen sensor to transfer the information related to hypoxia and oxidative stress into cells, activating intracellular defense mechanism against the hypoxic injuries.

Objective: To evaluate the feasibility and safety of applying enhanced recovery after surgery (ERAS) protocol in patients undergoing laparoscopic distal pancreatectomy. Methods: Data of 36 patients undergoing laparoscopic distal pancreatectomy from May 2016 to May 2017 in the First Affiliated Hospital, Zhejiang University School of Medicine were reviewed. The patients were divided into ERAS group (n=12) and control group (n=24). The patients in ERAS group received a series of enhanced recovery procedures, including multimodal analgesia, early off-bed activity and early oral food-taking, etc. Operation time, intraoperative blood loss, time to first flatus, postoperative complications, and length of postoperative hospital stay were evaluated. Results: There were no statistically significant differences in operation time and intraoperative blood loss between ERAS group and control group (all P>0.05). The time to first flatus and length of postoperative hospital stay were significantly shortened in ERAS group (all P < 0.05). The ERAS group had lower incidence of postoperative complications (41.7% vs. 66.7%), and the complications in ERAS group tended to be milder, but the differences failed to show statistical significance (all P>0.05). Conclusion: The ERAS protocol for laparoscopic distal pancreatectomy can significantly promote gastrointestinal function recovery and shorten postoperative hospital stay, and may reduce the incidence of postoperative complications.

Objective: To evaluate the clinical efficacy of modified lateral window for maxillary sinus floor augmentation (MSFA). Methods: Fifty-five patients who visited the Stomatology Hospital Affiliated to Zhejiang University School of Medicine between June 2012 and October 2014 were enrolled in the study. Patients underwent MSFA with Bio-Oss grafts based on modified access window. During the operation the vertical height of the bony window was reduced from 6-8 mm of conventional oval window to 4-5 mm of slot-shaped window. The sinus membrane was detached completely via the lateral access and large particle Bio-Oss graft was placed in the sub-mucosal space. The implant survival, graft height, graft volume and resorption rates were measured. Intra-op and post-op complications were recorded. Results: There were 86 implants inserted. The 2-4 year cumulative survival rates were 97.67% by implant-based analysis and 96.36% by patient-based analysis. The residual bone height was (4.7±2.6) mm and bone width was (8.4±2.7) mm. The bone height of implantation site immediately after operation was (16.1±2.5) mm and it was (16.2±2.2) mm at restoration. The bone heights at 1 and 2 years after operation were (14.9±2.5) mm and (13.6±2.6) mm, respectively. The graft height was (10.6±2.8) mm and the graft volume was (1569±745) mm³ immediately after operation. The resorption rate of graft height 6 months after operation was 3.79% and that of graft volume was 7.87%. The 1-year accumulative resorption rate of graft height was 6.63% and that of graft volume was 10.89%. The 2-year accumulative resorption rate of graft height was 7.58% and that of graft volume was 15.26%. Small membrane perforation during MSFA was observed in 5 cases and all were successfully repaired by a collagen Bio-Gide membrane. Conclusion: The modified lateral technique obtains high implant survival rate, excellent graft stability and low complication rate at 2-4 year clinical follow-up, indicating that it is a safe, predictable and minimally invasive surgical method for severe atrophic maxillary posterior dentition.

Objective: To evaluate the efficiency and safety of endoscopic trans-fistula drainage (ETFD) for gastroesophageal anastomotic fistula with para-fistula abscess after esophagectomy. Methods: Among 456 esophageal cancer patients receiving esophagectomy between February 2012 and February 2017 in Sir Run Run Shaw Hospital, 15 cases were diagnosed as gastroesophageal anastomotic fistula with para-fistula abscess after surgery. Seven cases received ETFD treatment (ETFD group), and 8 cases received conventional treatment (control group). Recovery of inflammatory markers and fistula, length of hospital stay after esophagectomy and total medical expenses were compared between ETFD group and control group. Results: All patients recovered in ETFD group. Time of white cell count returning to normal and decline of C-reactive protein, time of fistula healing and length of hospital stay after esophagectomy in ETFD group were significantly shorter than those of control group (all P < 0.05). And medical expenses in ETFD group was also lower (P < 0.05). Conclusion: ETFD is effective and safe for gastroesophageal anastomotic fistula with para-fistula abscess after esophagectomy.

Objective: To verify whether the enzymatic activity of kynureninase (KYNU) could be changed by the Arg188Gln (G/A) mutation. Methods: The total RNA of human hepatic tissue was extracted and the KYNU gene cDNA was amplified by RT-PCR. Primers were designed according to the sequences around the site Arg188Gln of KYNU gene and the Arg188Gln (G/A) mutant KYNU cDNA was generated by site-directed mutagenesis. Both the wild-type and mutant-type KYNU genes were subcloned into pcDNA vectors and the recombinant plasmids were constructed. After being transfected into human embryonic kidney 293 (HEK293) cells, the expression of KYNU recombinant plasmids were assessed by Western blot. The enzymatic activities of KYNU were detected by high performance liquid chromatography (HPLC). Results: The KYNU enzyme activities were expressed in both wild and mutant HEK293 cells. Michaelis constants (Km) of the wild and mutant KYNU were (9.833±0.513) μmol/L and (29.900±0.265) μmol/L, respectively (P < 0.05). The maximum velocities (Vmax) of the wild and mutant KYNU were (0.700±0.096) nmol·mg^-1·min^-1 and (0.084±0.003) nmol·mg^-1·min^-1, respectively (P < 0.05). Conclusion: Arg188Gln (G/A) mutation can decrease the enzymatic activity of KYNU.

Objective: To evaluate the feasibility of two types of segmented biodegradable esophageal stents in treatment of refractory benign esophagus strictures. Methods: Uncovered biodegradable segmented stent and fully-covered biodegradable segmented stent were implanted into the porcine esophagus (6 for each). Data on biodegradation, complications, and tissue reactions were compared between two groups. Results: All animals kept good general conditions; no death, decreased food intake, weight loss and malnutrition were observed. No perforation, ulcer, hemorrhage, stent migration and severe complications occurred. Stents degradation commenced at week 3. Stents structure breakage and complete stents absorption occurred at week 7-8 and week 9-10 in uncovered stents. While in fully-covered stents, stents structure breakage and complete stents absorption occurred at week 8-9 and week 10-11. Hyperplasia was prominent at week 1-3 and ameliorated at week 6 after stent implantation. A longer degradation period was present in fully-covered stents than in uncovered stents, while fully-covered stents induced tissue reactions at early stage were mild. Conclusions: The application of biodegradable esophageal stents with a segmented trunk in refractory benign esophagus strictures worth further investigation. The fully-covered stent has longer degradation period, which may be more suitable for clinical use.

Objective: To investigate the effect of exogenous L-Arg on the survival of extended perforator flap in rats. Methods: Sixteen male Sprague Dawley rats were randomly divided into L-Arg group (n=8) and control group(n=8). The extended dorsal three-vascular territory perforator flaps were made in rats. L-Arg (400 mg·kg^-1·d^-1) was injected intraperitoneally in L-Arg group 1d before operation, immediately and 1-7 d after operation, while the same volume of saline was injected intraperitoneally in control group at the same time points. The appearance and distribution of blood vessels were observed, and the flap survival areas were measured 7d after operation. The tissue samples were harvested from choke zone Ⅱ for histological study and the expression of vascular endothelial growth factor (VEGF) was detected by immunohistochemistry and Western blot, respectively. Results: After 7d, the clearer vascular structure and more new vessels in choke zone Ⅱ were observed in L-Arg group. The survival rate of flap in L-Arg group was (88.42±4.19)%, which was significantly higher than that in control group[(76.52±5.37)%, t=3.707, P < 0.01]. The microvessel density and caliber of choke zone Ⅱ in L-Arg group was (29.47±5.28)/mm² and(47.27±5.32)μm, which were significantly higher than those in control group (t=2.694 and 2.389, P < 0.05 or P < 0.01). The immunohistochemistry and Western blot showed that the expression of VEGF in choke zone Ⅱ of L-Arg group was significantly higher than that in control group (t=9.428 and -3.054, P < 0.05 or P < 0.01). Conclusion: Exogenous L-Arg can increase the survival rate of extended dorsal perforator skin flap through promoting vascularization and dilatation of vessels in choke zone Ⅱ in rats.

A middle-aged male patient with recurrent hypoglycemia was diagnosed as insulinoma according to the symptoms and the laboratory tests. Laboratory tests also showed low cortisol and adrenocortico-tropic hormone (ACTH) levels, therefore adrenal insufficiency was diagnosed and hydrocortisone was given. The hormone replacement therapy was terminated after surgery, and ACTH and cortisol levels returned to normal.

Enhanced recovery after surgery is a multimodal perioperative strategy according to the evidence-based medicine and multidisciplinary collaboration, aiming to improve the restoration of functional capacity after surgery by reducing surgical stress, optimal control of pain, early oral diet and early mobilization. Compared with other sub-specialty in general surgery, pancreatic surgery is characterized by complex disease, highly difficult procedure and more postoperative complications. Accordingly, pancreatic surgery shares a slow development in enhanced recovery after surgery. In this review, the feasibility, safety, application progress, prospect and controversy of enhanced recovery after surgery in pancreatic surgery are discussed.

Enhanced recovery after surgery (ERAS) has been widely used in perioperative optimization. As an important component of ERAS, rehabilitation medicine mainly focuses on perioperative physical fitness management, respiratory training, exercise training to reduce the incidence of postoperative pulmonary infection, improve gastrointestinal and cardiopulmonary function. This paper explains rehabilitation medicine for respiratory, musculoskeletal, cardiovascular and digestive systems during the perioperative period.