BACKGROUND:Periosteum is considered as a source of seed cels for cel therapydue toits biological features. OBJECTIVE:To seek the optimal way to isolate and culture rabbit periosteal cels and identify their biological features. METHODS:Rabbit periosteum on facies medialis tibiae was taken out under aseptic conditions. Periosteal cels isolated through the digestion of type II colagenase with the explants culture method were cultured in DMEM/F12 complete medium. Cel ultrastructure was observedunderan inverted microscope. Periosteal cel proliferation was determined bycel counting kit-8assay. Cel surface antigensCD90 and CD105 were determined using flow cytometry. Osteogenic andlipogenic induction mediums were applied to induce periosteal cels to differentiate into osteocytes and adipocytes, respectively. After 2 weeks of induction, cels were harvestedfor alizarin red staining and oil red O staining to assay the calciumnodules and lipid droplet. RESULTS AND CONCLUSION:The digestion of type II colagenase with the explants culture method shortened the period of primary cels culture and enhanced the survival rate, which causedhigher purity and stronger reproductive activity of harvested periosteal cels. Primary cultured periosteal cels grew in form of spindle spiral or paralel. Alizarin red andOil red O staining verified the multi-directional differentiation potentiality of periosteal cels. These findings suggest that the periosteal cels with high purity,strong reproductive activity,andmulti-directional differentiation potentialitycanbe harvested in short time using digestion of type II colagenase with the explants culture method.

Genetic bistable systems are a large class of important biological systems. Bistability, the capacity to achieve two distinct stable steady states in response to a set of external stimuli, arises within biological systems ranging from the ? phage switch in bacteria to cellular signal transduction pathways in mammalian cells. On the other hand, the increasing experimental evidence in the form of bimodal population distribution has indicated that noise plays a very key role in the switching of bistable systems. However, the physiological mechanism underling noise-induced switching behaviors has not been well explored yet. In the previous work, it has been showed that noise can induce coherent switch for a single genetic Toggle switch system. Here the influence of several kinds of noises (including intracellular and extracellular noises) on synchronized switch was investigated for a multicell gene toggle switch network system. It has been found that multiplicative noises resulting from fluctuations of either synthesis or degradation rates and the additive noise within each cell (they altogether are called as intracellular noises) all can induce the synchronized switch, and that there exists an optimal noise intensity such that the synchronized switch is optimally achieved and the amplification factor has the maximal value. On the other hand, the extracellular noises arising from the stochastic fluctuation of the cellular environment, not only brings about the synchronized switch, but also enhances it by suppressing intracellular fluctuations when the intracellular noises are not enough to induce the synchronized switch. Finally, the influence of the diffusive rate of signal molecules affected by noise on the dynamics of the multicellular system was also investigated, showing that the larger the diffusive rate, the better the synchronized switch and the larger the amplification factor.

Objective:To explore the relationship between the length of cervical spinous process and cervical motion and affected segment of cervical spondylotic myelopathy(CSM).Methods:Retrospective analysis was performed on 375 patients who underwent cervical surgical treatment due to single-segment cervical spondylotic myelopathy from January 2015 to January 2019. There were 200 males and 175 females, aged 50.72±9.39 (range 40 to 60) years. Several parameters, including the sagittal diameter of vertebral body, the sagittal diameter of cervical canal, the length of cervical spinous process, C 3-C 7 lordotic angle, range of motion (ROM) at C 3-C 7 and segmental ROM were measured via preoperative plain radiographs. All parameters were tested via Shapiro-Wilk method. Pearson correlation analyses was used to quantify the relationship between the lengths of C 3-C 7 spinous process and segmental ROMs. Receiver operating characteristic (ROC) curve was mapped to obtain the cut-off points according to the length of cervical spinous process which had significant differences. Patients were divided into two groups based on the cut-off points. χ2 test and t test were used to exclude the interference of age, gender and other anatomical factors and compare the differences in the affected segment of cervical spondylotic myelopathy between groups, so as to analyze the relationship between the length of cervical spinous process and affected segment of cervical spondylotic myelopathy. Results:There were significant differences of C 6 spinous process 27.82±6.01 mm and significantly negative correlation between the length of C 6 spinous process and the ROM at C 6,7 segment ( r=-0.338, P<0.001), while no significant correlations were found in other segments. ROC curves were mapped to obtain the cut-off points, and the cut-off point was 0.76. Group I: the ratio of the length of spinous process of C 6/C 7 (C 6/C 7 ratio, range 0.49 to 1.01) under 0.76, Group II: C 6/C 7 ratio more than 0.76. Compared with patients with longer-type C 6 spinous process (C 6/C 7 ratio ≥0.76), patients with shorter-type C 6 spinous process (C 6/C 7 ratio <0.76) had significantly bigger ROM at C 6,7 segment (10.11° vs 7.10°, P<0.001) and higher incidence of C 6,7 spinal cord compression ( χ2=16.642, P<0.001, OR=2.521), while differences in age, sex, sagittal diameters of vertebral body and spinal canal between two groups were not significant. Conclusion:The length of C 6 spinous process was significantly correlated with ROM at C 6,7 segment and the incidence of C 6,7 degenerative myelopathy. The length of C 6 spinous process can be considered as a predictor of development of C 6,7 degenerative myelopathy.

Objective To explore the alteration of brain pain functional areas in patients with functional dyspepsia (FD) irritable bowel syndrome (IBS) overlap by rectal balloon volume stimulation and magnetic resonance imaging (MRI) and the differences with IBS alone patients and healthy individuals were compared.Methods A total of 11 IBS alone patients,16 IBS overlapped with FD patients (IBS-FD) and 10 healthy controls were recruited.Sensory thresholds and visual analogue scale (VAS) were recorded during the rectal balloon air injection process. The changes of brain activated areas were analyzed by functional MRI (fMRI) when the rectum was stimulated at the volume of 50,100 and 150 ml.The data were analyzed by least significant difference (LSD) test.Results Under rectal volumetric stimulation,the sensory thresholds of IBS-FD group and IBS alone group were (53.14 ± 16.05) ml and (59.20 ± 20.55) ml and the difference was not statistically significant (LSD test,P＞0.05).There was no significant difference in VAS score between IBS alone group and IBS-FD group (LSD test,P＞0.05).Rectal stimulated under different volume,the results of fMRI indicated the activation of anterior cingulated cortex, dorsolateral prefrontal cortex,postporietal cortex,thalamus and insular cortex in both IBS alone group and IBS-FD group.And there was no significant difference in activated areas and intensity between IBS alone group and IBS-FD group (LSD test,P＞0.05).Conclusions There was no significant difference in activations of brain areas between IBS alone and IBS-FD patients under rectal volumetric stimulation. Under rectal volumetric stimulation,although symptoms overlapped,there was no evidence of the overlap of braingut axis and visceral hypersensitivity between IBS alone and IBS-FD.

Objective To investigate the role of expression of tall-like receptor 4 (TLR4) on medullary system cell (neutrophii, platelet) and endothelial cell surface in polymorphonuclear neutrophils (PMN) recruitment in rata with acute necrotizing pancreatitis. Methods C3H/He-J mice and C3H/He-N mice were divided into 4 groups by bone marrow transplantation: mosaic of medullary system cell TLR4-/- and endothelial cell TLR4 +/+ ; mosaic of medullary system cell TLR4 +/+ and endothelial cell TLR4-/-; mosaic of medullary system cell TLR4 +/+ and endothelial cell TLR4 -/-; mosaic of medullary system cell TLR4-/- and endothelial cell TLR4-/-, another control group was also established. ANP was induced in all these groups by intraperitoneal injection of cerulein and caudal vein injection of lipopolysaccharide. Serum amylase, pancreatic tissue naphthol AS-D chloroacetate esterase, MPO activity, TRL4 mRNA expression in peripheral blood granuloeyte was determined by RT-PCR, TRL4 protein expression in pancreatic tissue was measured by immunohistochemisty. Results Compared with that of control group, the levels of serum amylase in the 4 groups all significantly elevated and there was no difference among these 4 groups. Pathological scores of pancreas in the 4 groups were 5.52 ± 1.21, 5.18 ± 1.02, 2.03 ± 0. 82, 1.92 ± 0. 78, respectively; MPO activities were (1.834 ± 0. 170) U/g, (2. 596 ±0. 138) U/g, (0. 367 ±0. 018) U/g, (0. 202 ±0. 018) U/G, respectively; AS-D counts were 66.88 ± 2.17, 75.00 ± 2.43, 21.50 ± 2.38, 20.00 ± 2.19, respectively ; the expressions of TLR4mRNA of granular cell in peripheral blood were 0. 037 ± 1. 047 E-2, 1. 489 ± 8. 084 E-2, 1.470 ± 5. 210E-2, 0. 017 ± 6. 668 E-3, respectively; the 2 groups with endothelial eel1 TLR4 +/+ had strongly positive expression of TLR4 protein in vascular endothelial cell ; while the 2 groups with endothelial cell TLR4-/- had no expression; the pancreatic injuries, MPO activities, AS-D counts and TLR4 protein in the 2 groups with endothelial cell TLR4 +/+ were significantly higher than those in the 2 groups with endothelial cell TLR4-/-. Conclusions It was endothelial cell, not peripheral blood granulocyte, which played a key role in the process of neutrophil recruitment and pathological injure of ANP.

Objective To investigate the significance of expression of toll-like receptor on medullary system cell and endothelial cell surface in PMN recruiting in severe acute pancreatitis.Method In this study,26 C3H/He-J mice and 26 C3H/He-N mice were derided into 4 groups by bone marrow transplantation:group A, mosaic of medullary system cell TLR4-/- and endothelial cell TLR4+/+,group B, mosaic of medullary system cell TLR4+/+ and endothelial cell TLR4-/- ;group C, homozygote ofTLR4+/+ ;group D, homozygote of TLR4-/-. SAP was induced to all these mice. Amylase, TRIA of granular cell in peripheral blood,TRL4 expression in pancreas, Naphthol AS-D chloroacetate esterase, MPO activity were tested. Results Amylase level increased in all mice, there was no differences of the amylase level among groups, TRIA of granular cell in peripheral blood in group B and C was significant higher than A and D(P<0.05), TLR4 in pancreas were positive in group A and C, negative in group B and D, AS-D count and M PO activity in group A and C were higher than that in group B and D (P < 0. 05 ). Conclusions It was endothelial cell, not peripheral blood granulocyte, that plays a key role in recuiting neutrophils that triggers severe acute pancreatitis.

BACKGROUND:Periosteal cel s have been used in bone repair, but whether nucleus puplousus cel s co-cultured with autologous periosteal cel s can differentiate into osteoblasts in spinal fusion is rarely reported. OBJECTIVE:To isolate nucleus puplousus cel s and periosteal cel s so as to observe the osteogenic ability of nucleus puplousus cel s co-cultured with periosteal cel s or not. METHODS:Type II col agenase digestion method was used to isolate and purify nucleus pulposus cel s, which were confirmed by toluidine blue and immunohistochemical staining. Periosteal cel s were isolated histological y and cultured in complete medium, and cel surface antigens CD90, CD105 were identified by immunofluorescence staining. According to the experimental needs, the cel s were assigned into two groups. Nucleus pulposus cel s and periosteal cel s were co-cultured by osteogenic induction medium in the experimental group. Nucleus pulposus cel s in the control group were cultured alone in osteogenic induction medium. Cel morphology was observed by inverted microscopy, and cel proliferation was detected by cel counting kit-8. The osteogenic differentiation indexes of cel s in each group were measured using alkaline phosphatase staining, alizarin red staining, and type I col agen immunohistoehemical staining. The expression of osteopontin was tested by western blot assay. RESULTS AND CONCLUSION:CD105 and CD90 expressions of the periosteal cel s were positive. Nucleus puplousus cel s were positive for toluidine blue and col agen type II immunohistochemical staining. The proliferative ability of nucleus puplousus cel s was significantly higher in the experimental group than the control group at days 1, 3, 5, 7, 9. After 2 weeks of induction, the cel s were positive for alkaline phosphatase staining, alizarin red staining, and type I col agen immunohistoehemical staining, but the experimental group showed higher positive expressions than the control group (P<0.05). The expression of osteopontin was also higher in the experimental group than the control group. These findings indicate that nucleus puplousus cel s possess osteogenic ability, but have lower proliferative ability in vitro. After co-culture with periosteal cel s, the proliferative ability of nucleus puplousus cel s can be increased. Under osteogenic induction, nucleus puplousus cel s co-cultured with periosteal cel s have good compatibility and adhere with each other, which have stronger osteogenic ability than cel s cultured alone.

Objective To evaluate a new platelet function analyzer PL-11 with three major platelet function methods.Methods A randomized controlled trial was adopted.Totally 20 healthy young students from People's Liberation Army Medical School were enrolled in the study during July and August in 2013.Subjects were treated with aspirin or clopidogrel and the platelet function was measured by using of PL-11,as well as light transmittance aggregometry (LTA),VerifyNow and thromboelastography (TEG).Results When monitor aspirin response,correlations between arachidonic acid (AA) induced PL-11 and other methods were:LTA,0.614; VerifyNow,0.829; TEG,0.697,respectively.Biases between AA induced PL-1 1 and LTA were 1.94％ at baseline and 24.02％ during aspirin treatment.Cut-off value of aspirin response tested with AA induced PL-11 was 33.3％.When monitor clopidogrel response,correlations between adenosine piphosphate (ADP) induced PL-11 and other methods were:LTA,0.767; VerifyNow,0.851; TEG,0.608.Biases between ADP induced PL-11 and LTA were 3.01％ at baseline and 6.56％ during clopidogrel therapy.Cut-off value of clopidogrel response tested with ADP induced PL-1 1 was 66％.Conclusion Results of different platelet function testing methods were not equally effective.PL-11 has a high capability when monitoring short aspirin or clopidogrel response.

Objective To discuss The Clinical Significance of Eosinophil (EOS) in urosepsis. Methods A total of 99 patients of urosepsis in Department of Urology,Guangdong Provincial TCM Hospital from Mar. 2013 to Jul. 2016 were selected as research objects by retrospective analysis. The patients were classified into groupEOS= 0 andgroup EOS > 0,group PCT(procalcitonin)≥ 2 ng/mL andgroup PCT < 2 ng/mL,the differences of PCT concentration and percentage of EOS in two groups were analyzed comparatively. 99 patients of urosepsiswere also compared the difference of the percentage of EOS with another group including 100 patients of urinary tract infection (UTI) without Sepsis. Results The percentage of EOS was significantly decreased in 86.9%(86/99)of patients of urosepsis. The paired student t test show the percentage of EOS in two days after treatment,four days after treatment, before hospital discharge were higher than that before the treatment, the difference wassignificant (P < 0.05). The Independent-Sample Test show that the PCTconcentration in EOS = 0 group were higher than EOS > 0 group,the percentage of EOS in PCT≥2 ng/mL groupwere lower than PCT<2 ng/mL group,difference were significant(P<0.05). And The Independent-Sample Testalso showed that the percentage of EOS of the Urosepsis group was definitely lower than the UTI group without Sepsis. Difference was statistically significant. Concusions The percentage of EOS could be applied to assess the severity of urosepsis, monitor the disease progression and evaluate the infection control. The cost was lower than PCT in therapeuticprocess ofurosepsis.

BACKGROUND:Lower cervical anterior transpedicular screw technology combines the advantages of the anterior and posterior surgery;therefore, the pressure releasing and reconstruction problems can be solved via one time anterior surgery, whereas, the difficulty and risk of the operation are increased. However, the three-dimensional (3D) printing assembly navigation template improves the safety and accuracy of screw placement. OBJECTIVE:To explore the feasibility and accuracy of 3D printing assembly navigation template in lower cervical anterior transpedicular screws and compare it with free hand pedicle screw placement. METHODS:Lower cervical spine specimens of six adult (2 males, 4 females, average age 58.5 years old ranged from 53 to 64 years) corpses were equal y and randomly divided into two groups. Group A underwent free hand pedicle screw placement. Groups B (lower cervical anterior transpedicular screws assisted by personalized 3D printing combined navigation template):Three cadaveric lower cervical spines were examined using CT and data in DICOM format were recorded. After data were processed using software Mimics for 3D model reconstruction, computer-assisted design of optimum trajactory for lower cervical (C3-C7) anterior transpedicular screws placement was worked out and made into a dril template, where the surface was created as the inverse of anterior surface of cervical vertebra. The dril template was materialized in a 3D printing and used to place the screws. Subsequently, CT scan was performed to evaluate the screw orientation and acceptability. RESULTS AND CONCLUSION:(1) Thirty screws were inserted in Group A. The pedicle perforation was classified by CT, Grade 1:22 screws, Grade 2:6 screws, Grade 3:2 screws;insertion rate was acceptable (Grades 1-2):28 (93%). Thirty screws were inserted in Group B. The pedicle perforation was classified by CT, Grade 1:25 screws, Grade 2:4 screws, Grade 3:1 screw;insertion rate was acceptable (Grades 1-2):29 (97%). There were no statistical y significant differences in the rate of acceptable insertion and orientation between two groups (P>0.05). (2) These results suggested that 3D printing combined navigation template consisted with dril hole cap and screw hole, with functions of double direction. Compared with the traditional method, personalized 3D printing combined navigation template can be used simply.