Modulation of osteoclastogenesis through adrenomedullin receptors on osteoclast precursors: initiation of differentiation by asymmetric cell division.

Adrenomedullin (ADM), a member of the calcitonin family of peptides, is a potent vasodilator and was shown to have the ability to modulate bone metabolism. We have previously found a unique cell surface antigen (Kat1 antigen) expressed in rat osteoclasts, which is involved in the functional regulation of the calcitonin receptor (CTR). Cross-linking of cell surface Kat1 antigen with anti-Kat1 antigen monoclonal antibody (mAbKat1) stimulated osteoclast formation only under conditions suppressed by calcitonin. Here, we found that ADM provoked a significant stimulation in osteoclastogenesis only in the presence of calcitonin; a similar biological effect was seen with mAbKat1 in the bone marrow culture system. This stimulatory effect on osteoclastogenesis mediated by ADM was abolished by the addition of mAbKat1. 125I-labeled rat ADM (125I-ADM)-binding experiments involving micro-autoradiographic studies demonstrated that mononuclear precursors of osteoclasts abundantly expressed ADM receptors, and the specific binding of 125I-ADM was markedly inhibited by the addition of mAbKat1, suggesting a close relationship between the Kat1 antigen and the functional ADM receptors expressed on cells in the osteoclast lineage. ADM receptors were also detected in the osteoclast progenitor cells in the late mitotic phase, in which only one daughter cell of the dividing cell express ADM receptors, suggesting the semiconservative cell division of the osteoclast progenitors in the initiation of osteoclastogenesis. Messenger RNAs for the receptor activity-modifying-protein 1 (RAMP1) and calcitonin receptor-like receptor (CRLR) were expressed in cells in the osteoclast lineage; however, the expression of RAMP2 or RAMP3 was not detected in these cells. It is suggested that the Kat1 antigen is involved in the functional ADM receptor distinct from the general ADM receptor, consisting of CRLR and RAMP2 or RAMP3. Modulation of osteoclastogenesis through functional ADM receptors abundantly expressed on mononuclear osteoclast precursors is supposed to be important in the fine regulation of osteoclast differentiation in a specific osteotrophic hormonal condition with a high level of calcitonin in blood.

In vitro and in vivo detection of tunneling nanotubes in normal and pathological osteoclastogenesis involving osteoclast fusion.

Osteoclasts are multinucleated cells formed through specific recognition and fusion of mononuclear osteoclast precursors derived from hematopoietic stem cells. Detailed cellular events concerning cell fusion in osteoclast differentiation remain ambiguous. Tunneling nanotubes (TNTs), actin-based membrane structures, play an important role in intercellular communication between cells. We have previously reported the presence of TNTs in the fusion process of osteoclastogenesis. Here we analyzed morphological details of TNTs using scanning electron microscopy. The osteoclast precursor cell line RAW-D was stimulated to form osteoclast-like cells, and morphological details in the appearance of TNTs were extensively analyzed. Osteoclast-like cells could be classified into three types; early osteoclast precursors, late osteoclast precursors, and multinucleated osteoclast-like cells based on the morphological characteristics. TNTs were frequently observed among these three types of cells. TNTs could be classified into thin, medium, and thick TNTs based on the diameter and length. The shapes of TNTs were dynamically changed from thin to thick. Among them, medium TNTs were often observed between two remote cells, in which side branches attached to the culture substrates and beaded bulge-like structures were often observed. Cell-cell interaction through TNTs contributed to cell migration and rapid transport of information between cells. TNTs were shown to be involved in cell-cell fusion between osteoclast precursors and multinucleated osteoclast-like cells, in which movement of membrane vesicles and nuclei was observed. Formation of TNTs was also confirmed in primary cultures of osteoclasts. Furthermore, we have successfully detected TNTs formed between osteoclasts observed in the bone destruction sites of arthritic rats. Thus, formation of TNTs may be important for the differentiation of osteoclasts both in vitro and in vivo. TNTs could be one target cellular structure for the regulation of osteoclast differentiation and function in bone diseases.

Replacement Process of Carbonate Apatite by Alveolar Bone in a Rat Extraction Socket.

The objective of this study was to investigate a bone graft substitute containing carbonate apatite (CO3Ap) to analyze bone replacement and the state of bone formation in vitro and in vivo compared with autogenous bone (AB) or control. An osteoclast precursor cell line was cultured with AB or CO3Ap, and morphological analysis using scanning electron microscopy and a tartrate-resistant acid phosphatase activity assay were performed. The right maxillary first and second molars of Wistar rats were extracted and compensated by AB or CO3Ap granules. Following implantation, the bone formation state was evaluated after 3, 5, 7, 14, and 28 days of surgery by micro-computed tomography and immunohistostaining. The osteoclast-like cell morphology was typical with many cell protrusions in the AB and CO3Ap groups. Additionally, the number of osteoclast-like cells formed in the culture increased in each group; however, there was no significant difference between the AB and CO3Ap groups. Five days after tooth extraction, osteoclasts were observed near CO3Ap. The bone thickness in the CO3Ap group was significantly increased than that in the control group and the bone formation in the CO3Ap group increased by the same level as that in the AB group. CO3Ap is gradually absorbed by osteoclasts in the extraction socket and is easily replaced by alveolar bone. The process of bone replacement by osteoclasts is similar to that of autologous bone. By observing the process of bone replacement in more detail, it may be possible to gain a better understanding of the bone formation and control the amount of bone after surgery.

Construction and application evaluation of the information network platform for neonatal disease screening in Hubei Province / 公共卫生与预防医学

Objective To use the neonatal disease screening information platform to evaluate the effectiveness of screening for neonatal diseases in Hubei Province. Methods A total of 74 214 live births in the area under the jurisdiction of the Hubei Newborn Disease Screening Center and the newborns screened in the manual registration of disease screening in 2018 were selected as the control group. In addition, A total of 55 952 cases of live births in the same jurisdiction area in 2019 and neonatal data recorded in the neonatal disease screening system were included in the experimental group. The comparisons of the neonatal phenylketonuria, primary screening rate of congenital hypothyroidism, rescreening diagnosis rate and follow-up treatment between the two groups were carried out. Results The overall screening rate, the primary screening rate and rescreening diagnosis rate of the congenital hypothyroidism in the experimental group were higher than those in the control group, and the differences were statistically significant (P0.05). Conclusion The application of information network-based neonatal disease screening platform greatly improved work efficiency, reduced errors, enhanced screening rate and disease follow-up management capabilities, and thereby improved the management level of neonatal disease screening work.

CRISPR/Cas9 knockout plin1 enhances lipolysis in 3T3-L1 adipocytes / 生物工程学报

We used CRISPR/Cas9 to delete plin1 of 3T3-L1 preadipocyte, to observe its effect on lipolysis in adipocytes and to explore regulatory pathways. We cultured 3T3-L1 preadipocytes, and the plin1 knockout vectors were transfected by electroporation. Puromycin culture was used to screen successfully transfected adipocytes, and survival rates were observed after transfection. The optimized "cocktail" method was used to differentiate 3T3-L1 preadipocytes. The glycerol and triglyceride contents were determined by enzymatic methods. The changes in lipid droplet form and size were observed by Oil red O staining. The protein expression of PLIN1, PPARγ, Fsp27, and lipases was measured by Western blotting. RT-PCR was used to measure the expression of PLIN1 and lipases mRNA. After the adipocytes in the control group were induced to differentiate, the quantity of tiny lipid droplets was decreased, and the quantity of unilocular lipid droplets was increased and arranged in a circle around the nucleus. Compared with the control group, the volume of unilocular lipid droplets decreased, and the quantity of tiny lipid droplets increased after induction of adipocytes in the knockout group. The expression of PLIN1 mRNA and protein in the adipocytes was significantly inhibited (P<0.05); glycerol levels increased significantly (0.098 4±0.007 6), TG levels decreased significantly (0.031 0±0.005 3); mRNA and protein expression of HSL and ATGL increased (P<0.05); PPARγ and Fsp27 expression unchanged in adipocytes. The above results indicate that the knockout of plin1 enhances the lipolysis of 3T3-L1 adipocytes by exposing lipids in lipid droplets and up-regulating lipases effects.

PXR: a center of transcriptional regulation in cancer

Pregnane X receptor (PXR, NR1I2) is a prototypical member of the nuclear receptor superfamily. PXR can be activated by both endobiotics and xenobiotics. As a key xenobiotic receptor, the cellular function of PXR is mostly exerted by its binding to the regulatory gene sequences in a ligand-dependent manner. Classical downstream target genes of PXR participate in xenobiotic responses, such as detoxification, metabolism and inflammation. Emerging evidence also implicates PXR signaling in the processes of apoptosis, cell cycle arrest, proliferation, angiogenesis and oxidative stress, which are closely related to cancer. Here, we discussed, in addition to the characterization of PXR , the biological function and regulatory mechanism of PXR signaling in cancer, and its potential for the targeted prevention and therapeutics.

The significance of serum HE4 ,CA125 and ROMA indexes in diagnosis of ovarian cancer to different menopausal status / 西安交通大学学报(医学版)

Objective To evaluate the significance of HE4 ,CA125 and ROMA indexes in the diagnosis of ovarian cancer in different menopausal status women .Methods After the gynecological mass was taken for pathological diagnosis ,a total of 73 cases of ovarian cancer (ovarian cancer group ,divided into early stage and late stage) and 66 cases of gynecological benign tumor (benign tumor group ) were selected .57 healthy women were selected as controls (control group ) . All groups were further divided into non-menopausal and postmenopausal groups .Serum levels of HE4 and CA125 were measured by chemiluminescence immunoassay and the ROMA index was calculated . Results ① The serum levels of HE4 , CA125 and ROMA in ovarian cancer group were significantly higher than those in benign tumor group and healthy control group (P<0 .01) .② When benign tumor group and control group were taken as a reference , the receiver operating characteristic (ROC ) curve analysis showed that the area under the curve (AUC) of HE4 ,CA125 and ROMA in the non-menopausal group and the postmenopausal group was (0 .944 ,0 .822 ,0 .941 vs .0 .994 ,1 .000 ,1 .000) ,respectively .③ In non-menopausal group ,HE4 has higher specificity and positive predictive value .The best cut off value was 75 .92 pmol/mL ,the diagnostic sensitivity and specificity were 73 .1% and 100% .In postmenopausal group ,CA125 has higher sensitivity , the optimum cut off value was 57 .65 U/mL ,the diagnostic sensitivity and specificity were 100% and 95 .4% .④ In the postmenopausal group ,there was no significant difference in serum CA 125 level between the early and late ovarian cancer patients (P> 0 .05) .Conclusion In the premenopausal group HE4 has a better differential diagnostic value .In postmenopausal group ,the ROMA index could give consideration to both sensitivity and specificity ,and had higher diagnostic efficacy .It plays roles in the diagnosis of ovarian cancer to establish the appropriate reference interval of HE4 ,CA125 and ROMA indexes according to the different menopausal status .

Effects of application of citrate anticoagulation in bedside continuous blood purification of severe burn patients with sepsis / 中华烧伤杂志

Objective@#To investigate the effects of application of citrate anticoagulation in bedside continuous blood purification (CBP) of severe burn patients with sepsis, so as to provide reference for choosing anticoagulants in CBP of these patients.@*Methods@#Thirty severe burn patients with sepsis, conforming to the study criteria, were admitted to our burn intensive care unit from January 2014 to July 2017. Patients were divided into heparin group and citrate group according to computer randomization method, with 15 cases in each group. Patients in two groups all received bedside CBP treatment. Patients in heparin group used local heparin anticoagulation, while patients in citrate group used local citrate anticoagulation. Time of predicted single-time CBP treatment, time of single-time CBP treatment, time of accumulative CBP treatment, and rate of reaching the standard of CBP treatment time were counted. Changes of prothrombin time (PT), activated partial thromboplastin time (APTT), international normalized ratio (INR), fibrinogen, serum procalcitonin, and C-reactive protein (CRP) of patients before and after treatment were monitored. Hemorrhage in wounds, incision on trachea, and arteriovenous intubation point, and other complications during and after CBP treatment were observed. Data were processed with independent sample t test and chi-square test.@*Results@#(1) Time of predicted single-time CBP treatment of patients in the two groups was equal. Time of single-time CBP treatment and time of accumulative CBP treatment of patients in citrate group were longer than those in heparin group. Rate of reaching the standard of CBP treatment time of patients in citrate group was significantly higher than that in heparin group (χ2=16.655, P<0.01). (2) There was no statistically significant difference in PT, APTT, INR, fibrinogen, serum procalcitonin, and CRP of patients in the two groups before CBP treatment (t=0.203, -1.006, 0.203, 0.039, -1.591, -0.824, P>0.05). PT and APTT of patients in citrate group after CBP treatment were (14.2±1.6) and (45±7) s, respectively, significantly shorter than (15.5±1.4) and (53±6) s in heparin group (t=2.395, 3.321, P<0.05 or P<0.01). INR of patients in citrate group after CBP treatment was 1.13±0.12, significantly lower than 1.24±0.12 in heparin group (t=2.395, P<0.05). Fibrinogen of patients in citrate group after CBP treatment was (3.5±0.6) g/L, significantly higher than (3.0±0.6) g/L in heparin group (t=-2.427, P<0.05). Serum procalcitonin and CRP of patients in citrate group after CBP treatment were significantly lower than those in heparin group (t=2.520, 2.710, P<0.05). Decreased degree of serum procalcitonin and CRP of patients in citrate group after CBP treatment were (1.8±0.6) ng/mL and (143±69) mg/L, respectively, significantly higher than (0.9±0.6) ng/mL and (95±50) mg/L in heparin group (t=-4.033, -2.170, P<0.05 or P<0.01). (3) During CBP treatment, patients in heparin group experienced 21 times of exacerbation of wound hemorrhage and 10 times of new hemorrhage, including 2 times of hemorrhage at incision on trachea and 8 times of hemorrhage at arteriovenous intubation point. No exacerbation of hemorrhage or new hemorrhage happened in patients of citrate group. After CBP treatment, no electrolyte disturbance happened in patients of heparin group, but 1 patient in citrate group experienced hypocalcemia.@*Conclusions@#Application of citrate anticoagulation in bedside CBP of severe burn patients with sepsis shows light impact on systematic coagulation status, and can effectively decrease inflammation reaction of burn sepsis with low rate of hemorrhage.

A brief history of the discovery of PXR and CAR as xenobiotic receptors

The nuclear receptors pregnane X receptor (PXR) and constitutive androstane receptor (CAR) were cloned and/or established as xenobiotic receptors in 1998. Due to their activities in the transcriptional regulation of phase I and phase II enzymes as well as drug transporters, PXR and CAR have been defined as the master regulators of xenobiotic responses. The discovery of PXR and CAR provides the essential molecular basis by which drugs and other xenobiotic compounds regulate the expression of xenobiotic enzymes and transporters. This article is intended to provide a historical overview on the discovery of PXR and CAR as xenobiotic receptors.

Analysis of Pathogen Distribution and Drug Resistance of Burn Patients in Our Hospital from 2013 to 2015 / 中国药房

OBJECTIVE:To investigate the pathogen distribution and drug resistance of burn patients in our hospital,and to provide reference for rational use of antimicrobial agents in the clinic. METHODS:389 burn patients were selected from our hospi-tal during Jan. 2013 to Dec. 2015 by simple random sampling,and then analyzed retrospectively in respects of pathogen culture, identification and the results of sensitivity tests. RESULTS:678 clinical specimen were collected from 389 burn patients of our hos-pital,564 strains of pathogens were detected,with positive rate of 83.19%. Of 564 pathogens,there were 367 stains of Gram-nega-tive bacteria(65.07%),mainly including Pseudomonas aeruginosa(158 strains),Escherichia coli(67 strains),Acinetobacter bau-mannii(36 strains),Enterobacter cloacae(31 strains);there were 177 strains of Gram-positive bacteria(31.38%),mainly includ-ing Staphylococcus aureus (81 strains),Enterococcus (44 strains) and Staphylococcus epidermidis (26 strains);there were 20 strains of fungus(3.55%),mainly including Candida albicans(13 strains). There were 42 strains of ESBLs E. coli(62.69%)and 11 strains of ESBLs Klebsiella pneumoniae (40.74%). Gram-negative bacteria were highly resistant to aminoglycosides,β-lac-tamase,tetracycline and cephalosporin. P. aeruginosa was sensitive to colistin sulphate. E. coli,E. cloacae and K. pneumoniae were sensitive to imipenem,A. baumannii was sensitive to meropenem,and their resistant rates were 0. Gram-positive bacteria were highly resistant to many common antimicrobial agents;S. aureus was sensitive to vancomycin,S. epidermidis sensitive to van-comycin,teicoplanin and minocycline,and their resistant rates were 0. Resistant rates of C. albicans and C. tropicalis to amphoteri-cin B and 5-fluorocytosine were≤5%. CONCLUSIONS:Main pathogen of infection in burn patients of our hospital is Gram-nega-tive bacteria,mainly being P. aeruginosa. Producing enzymes and drug resistance of main pathogens are serious. It is necessary to standardize clinical application of antimicrobial agents and choose antimicrobial agents rationally according to etiological examina-tion and clinical symptoms.

Clinical Observation of Compound Xuelian Burning Wound Ointment in the Treatment ofⅡDegree Deep Burn / 中国药房

OBJECTIVE:To investigate the effects and safety of Compound xuelian burning wound ointment in the treatment ofⅡdegree deep burn. METHODS:Using consubstantial control method,2 burn wounds(Ⅱdegree deep burn,about 10 cm×10 cm) selected from symmetric or adjacent part of extremities in 80 patients withⅡdegree deep burn were divided into treatment area and control area. Treatment area was given Compound xuelian burning wound ointment,and control area was given Sulfadiazine silver cream. Healing time,degree of pain due to dressing change,the amount of infiltration liquid,wound infection rate,the incidence of scar formation and the incidence of ADR were observed. RESULTS:The healing time of wound in treatment area was signifi-cantly shorter than in control group [(18.7±3.6)d vs.(23.8±3.1)d],with statistical significance(P0.05). No obvious ADR was found in 80 patients. CONCLUSIONS:Compound xuelian burning wound ointment in the treatment of Ⅱdegree deep burn can relieve the degree of pain due to dressing change,control the amount of infiltration liquid,promote wound healing and reduce the occurrence of scar with good safety.

Her-2/neu gene and radiosensitivity / 中国癌症杂志

Her-2 receptor has already been identified as one of the most important biological markers of some malignant tumors,and also plays an important role in the biological behavior of those tumor cells.The over-expression of Her-2 receptor is associated with radioresistance of various tumor cells,and thus an antibody to Her-2/neu receptor can probably function as a radiosensitizer.In this review,we summarized some advances in the molecular mechanism and clinical aspects of the relationship between Her-2 and cellular radiosensitivity.