Development of high-concentration labeled colloidal gold immunochromatographic test strips for detecting african swine fever virus p30 protein antibodies.

African Swine Fever (ASF), caused by the African swine fever virus (ASFV), has inflicted significant economic losses on the pig industry in China. The key to mitigating its impact lies in accurate screening and strict biosecurity measures. In this regard, the development of colloidal gold immunochromatographic test strips (CGITS) has proven to be an effective method for detecting ASFV antibodies. These test strips are based on the ASFV p30 recombinant protein and corresponding monoclonal antibodies. The design of the test strip incorporates a high-concentration colloidal gold-labeled p30 recombinant protein as the detection sensor, utilizing Staphylococcal Protein A (SPA) as the test line (T line), and p30 monoclonal antibody as the control line (C line). The sensitivity and specificity of the test strip were evaluated after optimizing the labeling concentration, pH, and protein dosage. The research findings revealed that the optimal colloidal gold labeling concentration was 0.05 %, the optimal pH was 8.4, and the optimal protein dosage was 10 µg/mL. Under these conditions, the CGITS demonstrated a detection limit of 1:512 dilution of ASFV standard positive serum, without exhibiting cross-reactivity with antibodies against other viral pathogens. Furthermore, the test strips remained stable for up to 20 days when stored at 50 °C and 4 °C. Comparatively, the CGITS outperformed commercial ELISA kits, displaying a sensitivity of 90.9 % and a specificity of 96.2 %. Subsequently, 108 clinical sera were tested to assess its performance. The data showed that the coincidence rate between the CGITS and ELISA was 93.5 %. In conclusion, the rapid colloidal gold test strip provides an efficient and reliable screening tool for on-site clinical detection of ASF in China. Its accuracy, stability, and simplicity make it a valuable asset in combating the spread of ASF and limiting its impact on the pig industry.

Levels and dynamic changes of serum fibroblast growth factor 23 in hypophosphatemic rickets/osteomalacia.

BACKGROUND: Hypophosphatemic rickets/osteomalacia is a group of diseases characterised by defective mineralization of bone due to hypophosphatemia and low 1,25-dihydroxy vitamin D. To explore the role of fibroblast growth factor 23 (FGF-23) in the regulation of phosphate homeostasis, we measured the circulating concentrations of this growth factor in healthy individuals and in patients with hypophosphatemic rickets/osteomalacia. METHODS: Nineteen patients with hypophosphatemic rickets/osteomalacia were included in hypophosphatemic group (HP, 12 female and 7 male, mean age was 30 years), and 19 healthy age-matched individuals served as the control group. Full length FGF-23 fragments were measured by two-site enzyme-linked immunosorbent assay. RESULTS: Mean FGF-23 concentrations were significantly higher in the HP group ((87.4 +/- 43.6) pg/ml) compared with the control group ((19.2 +/- 6.16) pg/ml; P < 0.001). In 1 patient with tumour-induced osteomalacia, serum FGF-23 concentrations were 84.1 pg/ml; these concentrations were normalized 2 hours after a hemangiopericytoma resection (7.8 pg/ml). Subsequently, serum 1,25(OH)(2) vitamin D3 concentrations significantly increased from 21.3 pg/ml to 89.3 pg/ml, and serum phosphorus levels were normalized. CONCLUSIONS: Serum FGF-23 concentrations were markedly elevated in patients with hypophosphatemic rickets. FGF-23 plays an important role in the pathogenesis of hypophosphatemic rickets/osteomalacia.

[Theoretical study of geometrical structures and properties of several replaced mercapto molecules].

The geometrical structures of several Raman probe molecules were optimized using density functional theory (DFT) of the hybrid density functional B3LYP method and 6-311+ + G** basis set. Their energy gap, nucleus independent chemical shift (NICS), polarizability and vibration spectrum were studied. The theoretical results showed that: 4-MPY, MBA and PATP had planar structures, the angle of BDT between S-H and benzene ring plane was 20. 2 degrees, and the 4-MBT was 39. 6 degrees; they all have a strong aroma and a large value of polarization. The order of the average of molecular hyperpolarizability tensor was BDT > 4-MBT > 4-MBA > 4-MPY > PATP, and the trends of polarizability anisotropy invariant were 4-MBA > 4-MBT > BDT > PATP > 4-MPY.

Treatment of hypocalcemia caused by hypoparathyroidism or pseudohypoparathyroidism with domestic-made calcitriol: a prospective and self-controlled clinical trial.

BACKGROUND: Parathyroid hormone deficiency or resistance may cause hypocalcemia with related symptoms and signs. Lifelong treatment of calcium combined with vitamin D or its metabolites is always necessary for these patients. Here we reported a prospective and open-label trial to investigate the efficacy and safety of domestic-made calcitriol in treatment of hypocalcemia caused by hypoparathyroidism or pseudohypoparathyroidism. METHODS: Twenty-four patients with confirmed hypoparathyroidism or pseudohypoparathyroidism aged (36.5 +/- 11.0) years old were studied. Among them, 16 patients had idiopathic hypoparathyroidism, 2 had pseudohypoparathyroidism and 6 had hypoparathyroidism secondary to cervical surgery. Serum calcium levels were lower than 1.88 mmol/L. Oral calcitriol was administered twice or three times with elemental calcium 1.2 g per day. All patients were followed every 4 weeks throughout the 12-week period. Dose adjustments of calcitriol were based on serum and urinary calcium levels and symptoms of hypocalcemia. RESULTS: Twenty patients were included by the end of this study. Muscular weakness, cramps, extremity paresthesia, Chovestek's sign and Trousseau's sign were relieved in 76.9%, 100%, 94.4%, 93.3% and 78.9% of patients, respectively. Serum calcium, plasma ionized calcium and serum phosphorus levels were (1.54+/-0.25) mmol/L, (0.64+/-0.10) mmol/L and (2.00+/-0.46) mmol/L at baseline, and reached (2.20+/-0.20) mmol/L, (0.95+/-0.06) mmol/L and (1.68+/-0.25) mmol/L (P<0.01) at the 12th week of treatment, respectively. Eighty percent of patients were assessed as effective and 20% as partly effective. Three, four and eight patients had hypercalciuria at the 4th, 8th and 12th week of treatment, respectively, which were reduced by thiazide diuretics. The final dose of calcitriol was (1.09+/-0.50) microg/d. CONCLUSIONS: Calcitriol combined with calcium can be used in treatment of hypocalcemia caused by hypoparathyroidism or pseudohypoparathyroidism effectively and safely. Serum and urinary calcium levels should be monitored during the course of the therapy.

[The efficacy and safety of intravenous bisphosphonates in the treatment of primary hyperparathyroidism complicated by hypercalcemia crisis].

OBJECTIVE: To study the efficacy and adverse events of intravenous bisphosphonates in the treatment of patients of primary hyperparathyroidism (PHPT) complicated by hypercalcemia crisis. METHODS: From October 2003 to December 2007, 14 patients admitted into our hospital were diagnosed as PHPT complicated by hypercalcemia crisis, which was defined as a serum calcium concentration greater than 3.50 mmol/L. Of them, 6 cases had parathyroid adenoma, 1 had hyperplasia and 7 had parathyroid carcinoma. One of the intravenous bisphosphonates including pamidronate, ibandronate and zoledronic acid was given for 29 times in all the 14 cases. Serum calcium, parathyroid hormone, hematology, and other biochemical markers were monitored. Adverse events were recorded. RESULTS: After intravenous bisphosphonates, the serum total calcium (Ca) levels decreased from (3.85 +/- 0.50) mmol/L to (2.86 +/- 0.39) mmol/L in (1.4 +/- 0.6) days, and were kept below 3.50 mmol/L for (10.14 +/- 8.54) days. There was no significant difference of the magnitude of decrease in serum Ca levels among the patients using pamidronate, ibandronate or zoledronic acid. The change of serum Ca level was associated with the serum Ca level before treatment. The response to intravenous bisphosphonates evaluated by the decrease of serum total calcium levels was more significant in patients with parathyroid adenoma or hyperplasia than those with parathyroid carcinoma. The most common adverse event was pyrexia, which occurred 15 times (51.7%) and 75% of the pyrexia events occurred after the first infusion. Other manifestations included fatigue, flu-like symptom, myalgia, arthralgia and diarrhea with an incidence of 3.4% each (one event in the 29 times of treatment). There were 2 events (6.7%) with mild increase of serum creatinine concentration. CONCLUSION: Bisphosphonates can decrease serum total calcium levels in hypercalcemia crisis caused by PHPT effectively with mild adverse events.

[Association of calcium-sensing receptor gene polymorphism with serum calcium level in healthy young Han women in Beijing].

OBJECTIVE: To investigate the distribution of calcium-sensing receptor (CASR) gene polymorphisms in healthy young women of Han nationality in Beijing area and to explore the association of CASR genotypes and serum calcium, parathormone (PTH) level in healthy women. METHODS: 202 healthy young women aged (27 +/- 5) years of Han nationality in Beijing area were recruited in this study. Whole blood genome DNA was extracted with QIAGEN DNA extraction kit. A986S and G990R genotypes were determined with mutagenically separated polymerase chain reaction (MS-PCR) and PCR-restriction fragment length polymorphisms (RFLP), respectively. Serum calcium, albumin, phosphorus, PTH were determined. RESULTS: (1) There were A986S and G990R polymorphisms in healthy young women of Han nationality in Beijing. The frequencies of genotypes were as follows: SS absent, AA 95.0% and AS 5.0% for A986S, RR21.3%, GR51.0% and GG27.7% for G990R. Hardy-Weinberg equilibrium was evident. (2) In subjects carrying AA and AS genotype, serum Ca was (2.46 +/- 0.09) mmol/L vs (2.45 +/- 0.08) mmol/L, calcium adjusted by albumin was (2.31 +/- 0.10) mmol/L vs (2.30 +/- 0.09) mmol/L, serum P was (1.23 +/- 0.14) mmol/L vs (1.24 +/- 0.11) mmol/L, serum PTH was (41.6 +/- 18.6) ng/L vs (50.0 +/- 25.1) ng/L. There were no significant differences among them. (3) In subjects carrying GG, GR and RR genotype, serum Ca was (2.44 +/- 0.10), (2.46 +/- 0.08) and (2.48 +/- 0.08) mmol/L; calcium adjusted by albumin was (2.30 +/- 0.10), (2.32 +/- 0.09) and (2.32 +/- 0.10) mmol/L; serum P was (1.22 +/- 0.13), (1.24 +/- 0.15) and (1.20 +/- 0.15) mmol/L, serum PTH was (37.6 +/- 16.0), (42.1 +/- 20.2) and (45.9 +/- 18.1) ng/L. There were significant difference in serum Ca, calcium adjusted by albumin and PTH (P = 0.042, 0.020 and 0.014, respectively). CONCLUSIONS: (1) There are A986S and G990R polymorphisms in CASR gene in Han nationality in Beijing area. The frequencies of genotypes are as follows: SS absent, AA 95.0% and AS 5.0% for A986S, RR21.3%, GR51.0% and GG27.7% for G990R. Hardy-Weinberg equilibrium is evident. A and G alleles are more common. It is indicated that the distribution of CASR gene polymorphisms in Chinese is different from that in Caucasians. (2) G990R polymorphism is associated with serum calcium and PTH levels in healthy women. Subjects with R allele have higher levels of serum calcium and PTH. There is no correlation between A986S polymorphism and serum calcium or PTH.

[Skeletal biomechanical properties improvement in ovariectomized osteoporotic rats of teriparatide was retarded by alendronate].

OBJECTIVE: To implore the effects of teriparatide (PTH) and alendronate (Alen) on skeletal biomechanical properties of ovariectomized (OVX) osteoporotic rats. METHODS: 70 female Wistar rats of 6 months were randomly divided into 7 groups: (1) Baseline: sacrificed at baseline; (2) OVXb: sacrificed in 6 weeks after OVX; (3) Sham operation; (4) OVXe: sacrificed 14 weeks after OVX; (5) PTH: 40 microg.kg(-1).d(-1); (6) Alen: 100 microg.kg(-1).d(-1); (7) A + P: PTH 40 microg.kg(-1).d(-1) and Alen 100 microg.kg(-1).d(-1). In group (5)-(7), different medicines were injected 5 times per week from 6th to 14th week after OVX. The cancellous biomechanical properties were measured with indentation test and the cortical properties were investigated with three-point bending test. RESULTS: (1) The can load and can stiff of distal femur of OVXb were significantly lower than those of baseline (P < 0.01). It is indicated that osteoporotic rat models with impaired bone strength were established. (2) The can load and can stiff of distal femur of PTH [(36.3 +/- 9.2) N, (160.7 +/- 48.0) N/mm(2)], Alen [(42.7 +/- 13.0) N, (122.9 +/- 35.6) N/mm(2)] and A + P [(44.3 +/- 18.2) N, (105.2 +/- 58.4) N/mm(2)] groups were all higher than those of OVXe [(19.5 +/- 8.5) N, (83.2 +/- 37.7) N/mm(2), P < 0.001 or P < 0.01]. In femoral shaft, maximal load and elastic load in PTH, Alen and A + P groups were higher than those of OVXe (P < 0.001, P < 0.01 or P < 0.05). (3) Can stiff in distal femur was higher in PTH than that in Alen and A + P groups (P < 0.05). Elastic load, maximal load and energy absorption in femoral shaft were higher in PTH and A + P groups than those in Alen group (P < 0.05), and maximal stress was higher in PTH than that in A + P and Alen group (P < 0.05). CONCLUSION: PTH and Alen were effective in improving the skeletal mechanical properties of OVX rats, but improvement in Alen and A + P was not so good as that in PTH group. It is indicated that anabolic effects of PTH was retarded by Alen's significant inhibition of bone turnover, therefore, PTH and Alen should not be utilized simultaneously.

[Effects of teriparatide and alendronate on bone mineral density of osteoporotic rats].

OBJECTIVE: To investigate the effects of teriparatide (hPTH1-34, PTH) and alendronate (Alen) on bone turnover rate and bone mineral density (BMD) of ovariectomized (OVX) osteoporotic rats. METHODS: 70 female 6-month-old Wistar rats were randomly divided into 7 groups: (1) baseline group: killed immediately as baseline controls; (2) sham operation group: injected subcutaneously with normal saline (NS) as normal controls; (3) OVXb group: underwent ovarietomy (OVX) and killed 6 weeks after OVX as pre-therapeutic controls; (4) OVXe group: injected with NS subcutaneously and then sacrificed 14 weeks after OVX as controls by the end of treatment; (5) PTH group: PTH 40 microg.kg(-1).d(-1) was administered; (6) Alen group: Alen 100 microg.kg(-1).d(-1) was administered; (7) A + P group: PTH 40 microg.kg(-1).d(-1) and Alen 100microg.kg(-1).d(-1) were administered. In groups 4 approximately 7, different medicines were injected subcutaneously QD 5 times per week from the 6th week to the 14th week after OVX and then the rats were killed and their right femurs, lumbar vertebrae, and samples of blood and urine were collected. Absorptometry was used to measure the BMD of the right femur and lumbar vertebrae. The serum calcium, phosphate, creatinine, alanine transaminase, and alkaline phosphatase (ALP) activity were measured by automatic biochemical analysis. The bone resorption marker urine deoxypyridinoline/creatinine (UDpd/Cr) level was measured by enzyme-linked immuosorbent assay. RESULTS: Six weeks after OVX the ALP and UDpd/Cr levels in the OVXb group were 101 U/L +/- 59 U/L and (118 +/- 32) x 10(-6) respectively, both significantly higher than those of the baseline group (58 U/L +/- 10 U/L and (48 +/- 34) x 10(-6) respectively, both P < 0.01) and the BMD results of the OVXb group were all significantly lower than those in the baseline group (all P < 0.01), which indicated that an OVX osteoporotic rat model was established successfully. The ALP and UDpd/Cr levels of the Alen group were 61 U/L +/- 28 U/L and (17 +/- 39) x 10(-6), significantly lower than those of the PTH group 120 U/L +/- 36 U/L and (111 +/- 26) x 10(-6) respectively, both P < 0.01) and the UDpd/Cr levels of the A + P group were between those of the Alen group and those of the PTH group. The BMD levels of the femur and lumbar vertebrae of the PTH, Alen, and A + P groups were all significantly higher than those of the control groups (all P < 0.01), and were similar to or higher than those of the sham operation group without significant differences between the PTH and Alen groups. The BMD level of the lumber vertebrae of the A + P group were significantly higher than those in the PTH group (all P < 0.05), and the femoral BMD results of the A + P group were significantly higher than those in the PTH and Alen groups (all P < 0.01). CONCLUSION: PTH and Alen are all effective on osteoporosis. In particular, the combination of PTH and Alen is more effective on increasing the BMD level.

[Effects of different human parathyroid hormone 1-34 administration on SaoS-2 cells].

OBJECTIVE: To observe the effects of different human parathyroid hormone 1-34 (hPTH1-34) administration on SaoS-2 cells, and explore the mechanism of bone formation improvement. METHODS: Each cycle covered 48 h. SaoS-2 cells were continuously or intermittently stimulated by 50 ng/ml hPTH1-34 for 1, 3, 6, 12, and 24 h in each cycle. Total RNA was extracted by Trizol kit. Alkaline phosphatase (ALP), osteocalcin or bone Gla-containing protein (BGP) and cyclic adenosine monophosphate (cAMP) levels were measured by chemical method, radioimmunoassay and competitive protein binding method, respectively. c-fos gene expression was semi-quantified by reverse transcription polymerase chain reaction (RT-PCR). RESULTS: ALP level was time-dependently increased in 1, 3 and 6 h stimulation, especially in 3 and 6 h (compared with control, P < 0.01; P < 0.05 or P < 0.01 compared with continuous stimulation). The cAMP level was time-dependently increased in 3 and 6 h incubation (P < 0.05 compared with control and continuous stimulation). Intermittent hPTH1-34 stimulation had more effects on cAMP level than continous action (P < 0.001). hPTH1-34 intermittent stimulation of 1, 3, and 6 h enhanced c-fos gene expression time-dependently. CONCLUSIONS: Intermittent hPTH1-34 stimulation has a stronger effect on osteoblast than continuous action, especially in 3, 6 h in each cycle intermittent stimulation. The synchronous responses of c-fos, ALP and cAMP to hPTH1-34 suggest that hPTH1-34 affect Saos-2 cells through cAMP dependent protein kinase A (PKA) pathway and c-fos gene paly an important role.

[Changes of urinary deoxypyridinoline crosslink/creatinine in rats after ovariectomy and anti-osteoporotic intervention].

OBJECTIVE: To observe the changes of urinary deoxypyridinoline crosslink/creatinine (UDpd/Cr) in rats after OVX and intervention by estrogen and bisphosphonate and investigate the possible application of deoxypyridinoline in osteoporosis diagnosis and treatment. METHODS: 40 female 6-month-old virginal Wistar rats were divided into 5 groups, ovariectomized or sham ovariectomized. (1) Ovxb (n = 8): sacrificed at 6 weeks after OVX; (2) Sham (n = 8): sham ovariectomized; (3) Ovxe (n = 8): sacrificed at 14 weeks after OVX; (4) O + E (n = 9):OVX + 17 beta estradiol [20 micrograms/(kg.d) ih]; (5) O + C (n = 7):OVX + cimadronate [0.2 mg/(kg.d)]; Treatment started 6 weeks after OVX and lasted 8 weeks. Rats in group 2-5 were sacrificed at 14 weeks after OVX. Urinary and serum biochemical parameters were measured, pQCT scanning of femur, bone biomechanical test in femur were determined. RESULTS: OVX resulted in increasing of UDpd/Cr 133.3% (P < 0.01). The ratio of UCa/Cr also increased in OVX groups but without any significant compared with Sham (P > 0.05). UDpd/Cr were reduced by 54.6% and 51.8% (P < 0.01) in O + E, O + C group respectively compared with Ovxe. The significant negative correlationships were found between UDpd/Cr and bone mass, BMD and biomechanic characteristics. CONCLUSIONS: UDpd/Cr ratio is a sensitive bone resorption marker, a marked changes were observed when the rats ovariectomized or treated with estradiol and cimadronate. There were best correlation between UDpd/Cr and bone mineral density and bone biomechanic characteristics. It is fair to apply UDpd/Cr ratio for osteoporosis diagnosis and treatment.

[Evaluation of bone architecture and biomechanic properties by peripheral quantitative computed tomography in rats].

OBJECTIVES: To evaluate the value of peripheral quantitative computed tomography (pQCT) in measuring bone architecture and biomechanic properties. METHODS: 50 virgin female Wistar rats six months old were randomly divided into 4 groups: (1) 8 rats were killed as baseline group; (2) 8 rats underwent sham operation and then were killed 14 weeks after (sham operation group); (3) 16 rats underwent bilateral ovariectomy (OVX) without further intervention. Six and 14 weeks after the operation each 8 rats were killed (OVX group); and (4) 18 rats underwent OVX too. After the OVX 9 of the 18 rats were treated with 17beta-estradiol 20 micro g/kg/d IH and 9 rats were treated with estradiol valerate 800 micro g/kg/d po for 8 weeks respectively. Then the 18 rats were killed (OVX plus estrogen group, O + E group). The right tibiae of the rats were taken for histomorphometric analysis, and the right femora were prepared for pQCT scanning and bone biomechanical measurement with indentation test and three-point bending test. RESULTS: Histomorphometric analysis showed that the trabecular volume of proximal tibia (Cn-BV/TV) in the OVX group was 8.1 +/- 1.4%, significantly lower than that in the sham operation group (19.5 +/- 1.5%, P < 0.01). pQCT scanning showed that the femoral trabecular bone mineral content (Trab BMC) in the OVX group was 1.7 +/- 0.3 mg/mm, significantly lower than that in the sham operation group (3.2 +/- 0.5 mg/mm, P < 0.01) and the femoral trabecular bone mineral density (Trab BMD) in the OVX group was 158 +/- 32 mg/mm(3), significantly lower than that in the sham operation group (320 +/- 39 mg/mm(3), P < 0.01). The cancellous maximal load (Can load) of the distal shaft of femur in the OVX group was 12.5 +/- 2.5 N, significantly lower than that in the sham operation group (45.9 +/- 3.2 N, P < 0.01). The cancellous stiffness (Can Stiff) of the distal shaft of femur in the OVX group was 226 +/- 48 N/mm, significantly lower than that in the sham operation group (396 +/- 72 N/mm, P < 0.01). The Can load of O + E group was 21.8 +/- 3.7 N, significantly higher than that in the OVX group (P < 0.05). The Can Stiff of the O + E group was 382 +/- 54 N/mm, significantly higher than that in the OVX group (P < 0.05). There were no significant differences in cortical bone determined by pQCT as well as biomechanic properties in measured by three point test after OVX and estrogen treatment. A significant positive correlation was shown between Trab BMD and Cn-TV/BV and between Trab BMD and Tb N (r = 0.88 and 0.73, both P < 0.01). Similarly, both Trab BMC and Trab BMD of the femur were significantly correlated with the Can load and Can Stiff determined by indentation test (r = 0.47 - 0.68, all P < 0.01). There was also a significant correlation of parameters measured by pQCT in cortical bone with the maximal load and stiffness for the femur midshaft, and the best correlation was found between the maximal load of femur midshaft and Crt BMC and Crt A (both r = 0.76 and P < 0.01). CONCLUSION: The geometric, densitometric and mechanical properties in cortical and trabecular bones of rat can be well described by pQCT.

[Interaction effects between 1,25-dihydroxyvitamin D3 and transforming growth factor-beta1 on the proliferation and differentiation of human embryonic osteoblasts].

OBJECTIVE: To characterize the interaction effects between 1,25-dihydroxyvitamin D(3) [1,25(OH)(2)D(3)] and human transforming growth factor-beta1 (hTGF-beta1) on the proliferation and differentiation of human embryonic osteoblasts. METHODS: Human embryonic osteoblasts were cultured. 1,25(OH)(2)D(3), of the concentration of 50 nmol/L, hTGF-beta1 of different concentrations (1 x 10(-8) g/L, 1 x 10(-7) g/L, 1 x 10(-6) g/L, and 1 x 10(-5) g/L), and both hTGF-beta1 of different concentrations and 1,25(OH)(2)D(3) of the concentration of 50 nmol/L were added into the cultures with a stimulation time of 96 hours. 3-(4, 5-dimethylthiazol-2-yl) - 2, 5 - diphenyl - tetrazolium bromide (MTT) was added into the cultures. ELISA technique was used to measure the optical density values so as to observe cell's number and proliferation. Alkaline phosphatase (ALP) activity was tested by King's method. Osteocalcin (OC) was tested by radioimmunoassay. The mRNA expression level of SMAD proteins, the key proteins of the TGF-beta signal transduction passageway, was detected by reverse transcriptase polymerase chain reaction (RT-PCR). RESULTS: MTT staining showed an absorbance of 0.086 +/- 0.022 in the 1,25(OH)(2)D(3) group, significantly lower than that of the control group (0.124 +/- 0.031, P < 0.05). The ALP activity in the culture of 1,25(OH)(2)D(3) group, hTGF-beta1 groups, and combined 1,25 (OH)(2)D(3) and hTGF-beta1 groups were 1.3 - 2.0 times that of the control group (all P < 0.05). When the concentration of hTGF-beta1 is 1 x 10(-6) g/L, an interaction effect was found between 1,25(OH)(2)D(3) and hTGF-beta1 on the ALP activity of human embryonic osteoblasts. The OC value in cell medium was 5.3 micro g/L +/- 1.6 micro g/L and 5.4 micro g/L +/- 0.9 micro g/L respectively in the hTGF-beta1 1 x 10(-6) g/L and 1 x 10(-5) g/L combined with 1,25(OH)(2)D(3) (50 nmol/L) groups, significantly higher than that in the control group (P < 0.05), but no interaction effect was found between 1,25(OH)(2)D(3) and hTGF-beta1 on the OC production. hTGF-beta1 of different concentrations and hTGF-beta1 of different concentrations combined with 1,25(OH)(2)D(3) all increased the SMAD3 mRNA expression level in human embryonic osteoblasts (all P < 0.05). When the human embryonic osteoblasts were treated with 1,25(OH)(2)D(3) combined with 1 x 10(-6) g/L hTGF-beta1, its SMAD3 mRNA level reached the peak, about 6-fold that of the control group. 1,25(OH)(2)VD(3) and/or hTGF-beta1 did not influenced the SMAD4 mRNA expression. CONCLUSION: 1,25(OH)(2)D(3) inhibits the proliferation of human embryonic osteoblasts and increases the expression of osteoblastic markers, such as ALP and OC, thus promoting its differentiation. An interaction effect exists between 1,25(OH)(2)D(3) and hTGF-beta1 on the differentiation of human embryonic osteoblasts, which may be partly induced by SMA D(3) protein in the TGF-beta signal transduction passageway.

[The effects of continuous stimulation with different doses of parathyroid hormone 1-34 fragment on human SaoS-2 cells].

OBJECTIVE: To observe the effects of different doses of human parathyroid hormone fragment (hPTH)1-34 on SaoS cells and to explore the signal pathway and mechanism. METHODS: 5 x 10(4) cells/ml SaoS cells were seeded. Doses of 5, 50, 500 and 5 000 micro g/L hPTH1-34 were supplemented respectively. Total RNA was extracted by Trizol kit. Alkaline phosphatase (ALP), osteocalcin (BGP) and cAMP concentrations were measured by chemical, radioimmunoassay and competitive protein binding methods. c-fos gene expression level was semi-quantified by reverse transcriptase (RT)-PCR. RESULTS: ALP activity was higher in 500 micro g/L dose (P < 0.05 vs. control). BGP level was inhibited in 5 000 micro g/L dose (P < 0.05 vs. control and pretreatment). 50 and 500 micro g/L hPTH1-34 enhanced cAMP level significantly (P < 0.05 vs. control). No obvious increase of cAMP level was found in 5 000 micro g/L and 5 micro g/L dose groups (P > 0.05 vs. control and pretreatment). c-fos expression was higher in 50 and 500 micro g/L group. CONCLUSION: Different doses of hPTH1-34 exert distinct effects on osteoblasts. hPTH1-34 affects ALP and c-fos depending on protein kinase A signal pathway. hPTH1-34 exerts its action via regulation of osteoblasts by c-fos.