MicroRNA-497 suppress osteosarcoma by targeting MAPK/Erk pathway.

BACKGROUND: The aim of this study was to study the mechanism of miRNA-497 in the apoptosis of osteosarcoma cells. METHODS: MG-63 cells were divided into the three groups: NC, BL and miRNA groups, NC group were treated with nothing; BL group were transfected with blank vector; miRNA group were transfected with miRNA-497. Cell proliferation rate was detected by MTT method; Apoptosis rate was detected by flow cytometry and measuring the gene and protein expression of MAPK, Erk and P 21 by RT-PCR and Western blot. RESULTS: The cell proliferation rate of miRNA group was significantly lower compared to NC group and BL group (p < 0.05); while the apoptosis rate of miRNA group (32.17 ± 3.23 %) was significantly higher than that of NC group  (8.40 ± 1.78 %) and BL group (8.83 ± 0.99 %) (p < 0.05, respectively). Regarding the gene expression detection, we found that gene and protein expressions of MAPK, Erk and P21 of miRNA group were significantly different compared to NC and BL groups (p < 0.05, respectively). CONCLUSION: MiR-497 can activate P21 expression by inhibiting the expression of MAPK/Erk signaling pathway, thus promoting the apoptosis of osteosarcoma cells (Fig. 5, Ref. 18).

Overexpression of a glycine-rich protein gene in Lablab purpureus improves abiotic stress tolerance.

Glycine-rich protein (GRP) is involved in the response to abiotic and biotic stresses in plants. A novel GRP gene in Lablab purpureus has been identified. The cDNA of LpGRP was obtained from an SSH library constructed with root tissues of L. purpureus MEIDOU 2012 by waterholding for 10 days. The function of LpGRP was also evaluated in Arabidopsis. The cDNA of LpGRP has 555 bp and encodes a 184-amino acid protein. LpGRP was induced by drought and improved tolerance to abiotic stress. In LpGRP overexpressing Arabidopsis, the tolerance of transgenic seedlings to drought and salt was improved, and transgenic seeds showed insensitivity to both ABA and NaCl. The insensitivity to ABA indicated that there was crosstalk between LpGRP and ABA-responsive genes. These results indicated that LpGRP is a drought-responsive gene that can increase the drought and salt tolerance of Arabidopsis seedlings overexpressing LpGRP.

The association between the RAGE G82S polymorphism, sRAGE and chronic periodontitis in Taiwanese individuals with and without diabetes.

BACKGROUND AND OBJECTIVE: The present study investigated the association between the RAGE G82S polymorphism, the plasma levels of sRAGE and chronic periodontitis in subjects with and without diabetes mellitus (DM). MATERIAL AND METHODS: A total of 230 patients with DM and 264 non-DM participants were recruited for this study. Genotyping of the RAGE G82S polymorphism was accomplished using polymerase chain reaction-restriction fragment length polymorphism, and associations were analyzed with the chi-squared test and logistic regression analysis. RESULTS: In the non-DM group, the chi-squared test showed that the frequency distributions of the G82S polymorphism were significantly different between chronic periodontitis and non-chronic periodontitis subjects (χ(2) = 8.39, p = 0.02). A multivariate logistic regression model showed that the (G82S + S82S) genotypes were associated with a significantly increased risk of chronic periodontitis development compared to the G82G genotype (adjusted odds ratio = 2.06, 95% confidence interval: 1.08-4.07). In the DM group, there was no association between the G82S polymorphism and chronic periodontitis development when a multivariate logistic regression was performed. Plasma levels of sRAGE were significantly higher in subjects with the G82G genotype compared to those with the (G82S + S82S) genotypes in both the non-DM (856.6 ± 332.0 vs. 720.4 ± 311.4 pg/mL, p = 0.003) and DM groups (915.3 ± 497.1 vs. 603.5 ± 298.3 pg/mL, p < 0.0001). However, there was no difference in plasma sRAGE levels between chronic periodontitis and non-chronic periodontitis subjects in both the DM and non-DM groups. Moreover, when the subjects were further sub-divided by the G82S polymorphism, the difference in plasma levels of sRAGE between chronic periodontitis and non-chronic periodontitis subjects in the DM and non-DM groups remained statistically insignificant. CONCLUSIONS: The present study revealed that the RAGE G82S polymorphism was associated with chronic periodontitis in the non-DM group but not in the DM group. Our results also showed that the plasma levels of sRAGE were significantly higher in subjects with the RAGE G82G genotype, and this correlation was not affected by the presence of chronic periodontitis in the DM and non-DM groups.

HLA-A*31:01 and different types of carbamazepine-induced severe cutaneous adverse reactions: an international study and meta-analysis.

HLA-A*31:01 was reported to be associated with carbamazepine (CBZ)-induced severe cutaneous adverse reactions (SCAR), including drug reaction with eosinophilia and systemic symptoms (DRESS), Stevens-Johnson syndrome (SJS) and toxic epidermal necrolysis (TEN). We conducted an international study using consensus diagnosis criteria to enroll a total of 93 patients with CBZ-SCAR from Europe or Asia. We found that HLA-A*31:01 showed a significant association with CBZ-DRESS in Europeans (P<0.001; odds ratio (OR) (95% confidence interval (CI))=57.6 (11.0-340)), and the strong association was also found in Chinese (P<0.001; OR (95% CI)=23.0 (4.2-125)). However, HLA-A*31:01 had no association with CBZ-SJS/TEN in neither Chinese nor Europeans. By comparison, HLA-B*15:02 showed a strong association with CBZ-SJS/TEN in Chinese (P<0.001, OR (95% CI)=58.1 (17.6-192)). A meta-analysis of this and other published studies confirmed that in all populations, HLA-A*31:01 had an extremely strong association with CBZ-DRESS (P<0.001, a pooled OR (95% CI)=13.2 (8.4-20.8)), but a much weaker association with CBZ-SJS/TEN (P=0.01, OR (95% CI)=3.94 (1.4-11.5)). Our data revealed that HLA-A*31:01 is a specific predictor for CBZ-DRESS but not for CBZ-SJS/TEN. More studies are needed to investigate the genetic determinant of CBZ-SJS/TEN in Europeans. Considering the potential clinical utility, the cost-effectiveness of the combined HLA-A*31:01 and HLA-B*15:02 genetic test to prevent CBZ-SCAR in Chinese needs further investigation.

Mapping an aphid resistance gene in soybean [Glycine max (L.) Merr.] P746.

Soybean aphid (SA: Aphis glycines Matsumura) is one of the most serious pests of soybean [Glycine max (L.) Merr.] worldwide. A single dominant gene was found to control SA resistance in soybean line P746, which exhibits antibiosis resistance. This study aimed to define the location of the SA resistance gene in P746. A F2:3 mapping population, including 312 individuals, was created based on the cross of P746 and 'Dongnong 47'. Combined with bulked segregant analysis, all of the 1015 simple sequence repeats (SSR) from the soybean consensus map were used to locate the tentative genomic region of the SA resistance gene in P746. The effort resulted in the mapping of R_P746, the SA resistance gene in P746, and was flanked on either side by Satt334 and Satt335 on chromosome 13. By chromosome walking with SSRs from BARCSOYSSR_1.0, R_P746 was mapped between BARCSOYSSR_13_1278 and BARCSOYSSR_13_1363, with distances of 4.2 and 2.6 cM, respectively. The results indicate that R_P746 might be different to the SA resistance genes previously reported. The markers that are closely linked to R_P746 are expected to be useful for marker assisted selection in future soybean aphid resistance breeding programs.

Identification of a hidden outbreak due to the spread of a VIM-3-producing, extensive drug-resistant Pseudomonas aeruginosa (XDRPA) clone at a regional hospital in Taiwan.

A review of the annual prevalence of Pseudomonas aeruginosa at a regional hospital in Taiwan revealed a significant increase in the incidence of extensive drug-resistant P. aeruginosa (XDRPA) from 2∙1% in 2003 to 5∙8% in 2007. The first XDRPA isolate was recovered in 2001 from the emergency ward. The widespread dissemination of XDRPA isolates to more than 10 other wards was discovered the following year. Six pulsotypes of 67 XDRPA isolates from 2006 onwards were identified and 91% were a single strain, suggesting the existence of a hidden outbreak. Prior to the recognition of the outbreak, the majority of cases were not considered to be healthcare-associated infections until molecular evidence was provided. A cohort measure was launched by the infection control practitioners that effectively controlled the outbreak. Patients with XDRPA were mostly referred from neighbouring long-term care facilities, which may have been the reservoir of the XDRPA clone.

Effects of different contacting pressure on the transfer function between finger photoplethysmographic and radial blood pressure waveforms.

BACKGROUND: The blood pressure (BP) waveform is suggested to reflect the whole-body blood supply distribution, but its non-invasive assessment is not sufficiently user friendly for practical applications. The present authors studied the correlation between BP and photoplethysmography (PPG) waveforms, with the aim of determining the optimal range for contact pressure stimulation (PS) to produce a reliable transfer function in their harmonic parameters. METHODS: Finger PPG and radial-artery BP signals were measured simultaneously and noninvasively on healthy volunteers (n = 45). PS of 0-200 mmHg was applied to the finger, and 1 min data sequences were recorded. In frequency-domain analysis, linear regression was applied to the calculated amplitude ratios or the first five harmonics between BP and PPG waveforms. RESULTS: In the 60 mmHg-PS group, the BP-PPG regression of amplitude ratios was highest, and the agreements between them were also the best verified by Bland-Altman analysis. CONCLUSION: In the present study, frequency-domain analysis was performed to study the correlation between BP and PPG waveforms. Differences in pressure-induced-vasodilation responses underlie the different BP-PPG waveform correlations obtained by applying different PSs. The non-invasively derived PPG parameters might help to provide an easier method to acquire the radial-artery BP waveform, and hence broaden the application of BP waveform analysis.

Long-term outcome and short-term survival of patients with systemic lupus erythematosus after bacteraemia episodes: 6-yr follow-up.

OBJECTIVE: To describe the nature of bacteraemia in SLE patients and determine the short-term survival and long-term outcome of these patients. METHODS: Analysis of the medical records of 1442 SLE patients who were regularly followed up in a tertiary teaching medical centre from 2000 to 2005. RESULTS: Among 1442 SLE patients, 240 patients (17%) developed at least one episode of bacteraemia, corresponding to an incidence of 92.7 cases/1000 hospital admissions. Since SLE diagnosis, the overall survival of our patients was 92% at 5 yrs, 86% at 10 yrs and 79% at 15 yrs. However, after one episode of bacteraemia, the survival decreased to 76% at 30 days and 67% at 360 days. Of the 336 episodes of bacteraemia, 167 were community-acquired (49.7%) and 169 were nosocomial (50.3%). Staphylococcus aureus was the leading cause of Gram-positive bacteraemia. Among Gram-negative bacteria, non-typhoidal Salmonella and Escherichia coli were the most common species. Community-acquired Salmonella and Streptococcus bacteraemia were more common than nosocomial infections. Klebsiella and Acinetobacter spp. were significantly more responsible for nosocomial than community-acquired bacteraemia. Patients infected with Acinetobacter, Klebsiella or Pseudomonas had lower probabilities of 14-day survival (71.4, 55.6, 42.9%, respectively). CONCLUSIONS: Among SLE patients, an episode of bacteraemia was associated with an unfavourable long-term outcome. The bacterial species significantly influenced short-term survival. Therefore, when empiric antibiotic therapy is initiated in SLE patients who are suspected of bacteraemia, we suggest use of antibiotics that are effective against Pseudomonas, Klebsiella, Acinetobacter, S. aureus, and E. coli.

The levels of volatile sulfur compounds in mouth air from patients with chronic periodontitis.

BACKGROUND AND OBJECTIVE: Volatile sulfur compounds may be the main source of oral malodor. The aim of this study was to clarify the relationship between periodontal parameters and volatile sulfur compounds and to evaluate the improvement of several halitosis-related outcomes by tongue scraping, nonsurgical periodontal treatment (including oral hygiene instruction) and oral hygiene instruction/chlorhexidine + cetyl pyridinium gargling. MATERIAL AND METHODS: Seventy-two chronic periodontitis patients with heavy tongue coating were assessed for oral malodor and periodontal status. Oral malodor was evaluated by measuring the levels of volatile sulfur compounds using OralChroma and the organoleptic test score. Thirty participants were selected for the subsequent experiments: tongue scraping; nonsurgical periodontal treatment; and oral hygiene instruction/chlorhexidine + cetyl pyridinium gargling. Twenty-five participants completed all experimental stages. RESULTS: Significant correlations were observed between the organoleptic test score and hydrogen sulfide (H2S), methyl mercapton (CH3SH), tongue coating score and volatile sulfur compounds, which was also significantly correlated with bleeding on probing percentage and tongue coating score. Tongue scraping significantly reduced the levels of volatile sulfur compounds. Further reduction of volatile sulfur compounds after nonsurgical periodontal treatment and oral hygiene instruction/chlorhexidine + cetyl pyridinium gargling were noted compared with baseline. CONCLUSION: Volatile sulfur compounds, with H2S and CH3SH as the main components, in mouth air are the prominent elements of malodor. Volatile sulfur compounds were decreased by more than 50% after tongue scraping. Nonsurgical periodontal treatment and oral hygiene instruction/chlorhexidine + cetyl pyridinium gargling maintained a significantly lower level of malodor compared with baseline.

Clostridium innocuum is a vancomycin-resistant pathogen that may cause antibiotic-associated diarrhoea.

OBJECTIVES: Clostridium innocuum can cause extraintestinal infection in patients with underlying diseases. The role of C. innocuum in antibiotic-associated diarrhoea (AAD) remains unknown. METHODS: Clinical information of 103 patients from whom C. innocuum was isolated was reviewed. We carried out cellular and animal experiments to examine the pathogenic potential of C. innocuum in AAD. RESULTS: Eighty-eight per cent (91/103) of the 103 patients received antibiotics within 2 weeks of diarrhoea onset. Patients were further classified into two groups, severe colitis and diarrhoea, according to clinical severity level. The mortality rate was 13.6% (14/103) among the patients from whom C. innocuum was isolated. The lowest concentrations at which 90% of the isolates were inhibited for metronidazole and vancomycin were 0.5 and 16 mg/L, respectively. All isolates tested were susceptible to metronidazole but resistant to vancomycin. Nineteen randomly selected isolates (ten from severe colitis group, nine from diarrhoea group) were subjected to further in vitro cellular examinations. The level of cytotoxicity to Vero cells was significantly higher in isolates from the severe colitis group at both 24 and 48 hours after inoculation (24 and 48 hours, p 0.042 and 0.033, respectively). We observed apoptotic changes that subsequently led to cell death in C. innocuum-infected Vero cells. Tissue damages, necrotic changes and oedema were observed in the mouse ileal loop infected by C. innocuum. CONCLUSIONS: Vancomycin-resistant C. innocuum may play a potential role as a causative agent of AAD. The clinical manifestations of AAD caused by C. innocuum were diarrhoea or severe colitis, including pseudomembranous colitis.

Clostridium innocuum is a significant vancomycin-resistant pathogen for extraintestinal clostridial infection.

OBJECTIVES: Extra-intestinal clostridial infection (EICI) is rare but can be fatal. Traditional phenotypic methods can only assign many of the Clostridium species to the genus level. METHODS: A total of 376 non-repetitive Clostridium isolates from sterile sites were collected and subjected to matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) Biotyper analysis and 16S rRNA sequencing. Antimicrobial susceptibility was determined, and clinical characteristics of the patients were assessed. Clostridium innocuum isolates were characterized by genome sequencing and genotyping. We used molecular and cellular methods to explore the virulence and resistance mechanisms of C.innocuum. RESULTS: Clostridium innocuum was the second most common species to cause EICI, only next to Clostridium perfringens. All Clostridium isolates showed susceptibility to clindamycin, metronidazole, penicillin, piperacillin and ampicillin-sulbatam, while C. innocuum isolates were invariably resistant to vancomycin. Among 24 patients with EICI caused by C. innocuum, two (8.3%) had diarrhoea, three (12.5%) had soft-tissue infection, six (25%) had appendicitis and four (16.7%) each had shock and gastrointestinal perforation. The 30-day mortality was 16.7%. The C. innocuum isolated from different sites could not be separated from one another by genotyping. No known toxin genes were identified in the genome of C. innocuum but the species expressed cytotoxicity to epithelial cells. d-Alanine-d-alanine ligase, alanine racemase and d-alanyl-d-alanine carboxypeptidase are three main genes responsible for vancomycin resistance in C. innocuum. CONCLUSIONS: Vancomycin-resistant C. innocuum is a previously unrecognized, yet prominent, cause for EICI. Genome analysis showed that the species could carry a lipopolysaccharide-like structure that is associated with cytotoxicity to cells in vitro.

Correlations of androgen deficiency with clinical symptoms in Taiwanese males.

The purpose of this study was to elucidate correlations between different biochemical measurements of androgen deficiency and clinical symptoms in male residents of Taiwan. An investigation of the serum biochemical markers for androgen deficiency in 650 males, including total testosterone, calculated free testosterone, and bioavailable testosterone, was conducted. Measurements of clinical symptoms were obtained using a questionnaire of the androgen deficiency in the aging male (ADAM) by St Louis University (SLQ). Correlations among the biochemical markers, correlations of the biochemical markers and age, and relationships between the biochemical markers and the SLQ were evaluated. The sensitivity and specificity of the SLQ were determined. Bioavailable and calculated free testosterone correlated better with age than did total testosterone. Eighty percent of the men had a positive SLQ, and 20% had a negative SLQ. The percentage of positive SLQ results increased with age. No statistically significant difference was noted between the biochemical markers of bioavailable and calculated free testosterone levels and the SLQ status except for men aged over 70 years. The SLQ in this study showed an acceptable sensitivity of about 80%, but the specificity was poor (about 20%). In conclusion, bioavailable testosterone and calculated free testosterone were more-closely correlated with age and may be better biochemical markers for androgen deficiency. SLQ might not be a suitable single measurement for androgen deficiency and should be used together with biochemical markers.

Anomalies in cell wall turnover associated with the growth temperature of Bacillus subtilis.

Cell wall turnover appeared to be anomalously fast in Bacillus subtilis when the cells were grown at temperatures below 29 degrees C. Turnover rates k(generation-1), of exponential cultures at 25 degrees were approximately double those of cells grown at 37 degrees C. When autolysin levels were assayed in cell walls, it was found that the enzyme activities were constant between 25 degrees C and 40 degrees C, suggesting that there was no greater synthesis of autolysin at the lower temperature. Analyses of walls for individual components, extent of aminosugar substitution and extent of crosslinking, did not reveal significant differences between samples obtained from 25 degrees C or 37 degrees C cultures. The N-acetylmuramoyl-L-alanine amidase was stable over the temperature range studied. Lysis of cells, induced by carbonylcyanide-m-chlorophenylhydrazone, occurred at a faster rate for cells obtained at 25 degrees C than for cells obtained at 37 degrees C. In addition, the lysis of cells by hen egg white lysozyme was slightly faster when the cells were obtained from 25 degrees C cultures than from 37 degrees C cultures. It is possible the autolysin(s) responsible for cell wall turnover are cold-activated.

Isolation of Salmonella enterica serotype choleraesuis resistant to ceftriaxone and ciprofloxacin.

Salmonella enterica serotype choleraesuis (S choleraesuis) usually causes systemic infections in man that need antimicrobial treatment. We isolated a strain of S choleraesuis that was resistant to ceftriaxone and ciprofloxacin from a patient with sepsis. Ciprofloxacin resistance was associated with mutations in gyrA and parC, whereas the ampC gene (bla(CMY-2)), responsible for ceftriaxone resistance, was carried by a transposon-like mobile element. This element was found inserted into finQ of a potentially transmissible 140 kb plasmid, with an 8 bp direct repeat flanking the junction regions. The appearance of this resistant S choleraesuis is a serious threat to public health, and thus constant surveillance is warranted.

Interspecies comparison of renal cortical receptors for parathyroid hormone and parathyroid hormone-related protein.

Parathyroid hormone (PTH) and PTH-related proteins (PTHrP) interact with a common receptor in rat bone cells and in canine renal membranes with similar affinity, but PTHrP are substantially less potent than PTH in stimulating adenylate cyclase in canine renal membranes; in contrast, PTH and PTHrP are equipotent in stimulating adenylate cyclase in rat bone cells. This discrepancy has been largely viewed as reflecting differences in the relative efficiency of signal transduction of PTHrP between bone and kidney assay systems. To test the alternative (but not mutually exclusive) hypothesis that these differences could reflect interspecies differences in PTH receptors, we have characterized the bioactivity of amino-terminal PTHrP and PTH in rat and human renal cortical membranes (RCM) and compared them to results we previously reported in canine RCM. The stability of PTH and PTHrP peptides under binding and adenylate cyclase assay conditions was greater than 80% for each species. Competitive inhibition of [125I](Tyr36)hPTHrP-(1-36)NH2 binding to rat RCM by bPTH-(1-34) and (Tyr36)hPTHrP-(1-36)NH2 yielded nearly identical binding dissociation constants (3.7 and 3.6 nM, respectively), and binding to human RCM demonstrated slightly greater potency for PTHrP (0.5 nM) than for PTH (0.9 nM). Similarly, adenylate cyclase stimulating activity was equivalent for the two peptides in rat RCM, but PTHrP was twofold more potent than PTH in human RCM. Covalent photoaffinity labeling of protease-protected rat RCM yielded an apparent 80 kD receptor protein, and cross-linking of human RCM labeled an 85 kD receptor, indistinguishable in size from the canine renal PTH receptor. We conclude that rat, canine, and human renal cortical PTH receptors exhibit species specificity. The previously observed differences between rat bone cells and canine renal membranes in the efficiency of signal transduction by PTHrP may be explained, at least in part, by these species differences.

Immunoaffinity purification of parathyroid hormone-related protein from bovine milk and human keratinocyte-conditioned medium.

Parathyroid hormone-related proteins (PRHrP) are a novel family of proteins that appear to be responsible for humoral hypercalcemia of malignancy. Although PTHrP derived from human tumors have been purified and their N-terminal amino acid sequence determined, and although the structure of the PTHrP gene and its alternatively spliced mRNA transcripts have been defined, the secretory and circulating form(s) of the protein are unknown. Purification of PTHrP in the past has been difficult, requiring multiple chromatographic steps and months or years to complete. To define naturally occurring PTHrP species we have developed a rapid and efficient immunoaffinity purification method. Bovine milk (250 ml) and human keratinocyte-conditioned medium (3000 ml) were affinity purified using a 300 microliters affinity-purified polyclonal anti-PTHrP-(1-36) antibody column and a single RP-HPLC step. Purification required only 7-10 days and yielded a 3-4% recovery. Quantities of PTHrP sufficient for silver-stained SDS-PAGE, Western analysis, and N-terminal amino acid sequence were obtained. In contrast to conventional purification schemes, affinity purification of PTHrP is rapid and efficient and can be applied to biologic samples that contain PTHrP in low abundance. These methods can be applied to the purification and characterization of the as yet undefined secretory and circulating forms of PTHrP.

Co-purification of transforming growth factor beta-like activity with PTH-like and bone-resorbing activities from a tumor associated with humoral hypercalcemia of malignancy.

Humoral hypercalcemia of malignancy (HHM) is caused by a circulating bone-resorbing factor or factors. Suggestions as to the nature of this factor include PTH-like proteins, transforming growth factors, and bone-resorbing factors distinct from either of the first two classes of polypeptides. We investigated the occurrence of these three activities in a highly purified extract of the H-500 Leydig cell tumor which causes HHM when implanted into Fisher rats. PTH-like adenylate cyclase-stimulating activity (ACSA) was extracted from tumor tissue by sequential treatment with urea/HCl and ethanol/NaCl. Tumor extract was further purified by hydrophobic-interaction, gel-filtration, and reverse-phase HPLC steps to a specific activity of 1038 ng eq bPTH(1-34)/mg protein. Only the fraction pool containing ACSA demonstrated significant bone-resorbing (1.78-fold over basal) and transforming growth factor activity (epidermal growth factor (EGF)-dependent colony formation in soft agar suspension by NRK-49F indicator cells). A subsequent reverse-phase HPLC step produced material which contained both ACSA and transforming growth factor beta (TGF beta)-like activity in a single fraction. Whether the responsible mediator in this animal model has TGF beta-like properties as well as PTH-like and bone-resorbing activity remains to be determined.

Glycosylation of parathyroid hormone-related peptide secreted by human epidermal keratinocytes.

While the gene and mRNA transcripts encoding PTH-related peptide (PTHrP) have been well characterized, the actual secretory form(s) of the peptide is unknown. Accordingly, synthetic and recombinant PTHrPs employed to date for biological and immunological characterization have necessarily been of arbitrary lengths. No prior evidence for glycosylation of PTHrPs has been described. To define the naturally occurring form(s) of this peptide secreted by human epidermal keratinocytes, we have affinity purified, using an anti-PTHrP-(1-36) antibody column, human PTHrP secreted under conditions of protease protection. Human keratinocyte-conditioned medium collected without measures to protect against proteolytic degradation contains multiple PTHrP immunoreactive and bioactive species. In contrast, under conditions of protease protection, human keratinocyte-conditioned medium contains a single 18,000 mol wt (Mr) form of the peptide. In contrast to recombinant and synthetic PTHrPs, which migrate as distinct, well focussed bands on sodium dodecyl sulfate-polyacrylamide gel electrophoresis, this 18,000 Mr PTHrP displays the broad electrophoretic profile of a glycoprotein. Treatment of this peptide with trifluoromethanesulfonic acid, an agent that deglycosylates both O- and N-linked saccharides from their core proteins, shifted the Mr of the protein to approximately 10,000. In contrast, exposure of recombinant PTHrP-(1-141) to the same agent results in no change in electrophoretic mobility. These studies indicate that the 18,000 Mr species of PTHrP secreted by human epidermal keratinocytes is a glycoprotein.

Skin-derived fibroblasts respond to human parathyroid hormone-like adenylate cyclase-stimulating proteins.

Human tumors and keratinocyte-conditioned medium contain PTH-like adenylate cyclase-stimulating proteins. Human dermal fibroblasts have receptors that recognize PTH and a factor associated with humoral hypercalcemia of malignancy in rats. We examined 10 human dermal fibroblast lines for an adenylate cyclase response to PTH. Six of 10 lines tested displayed a definite response (2.4- to 3.8-fold over basal) to 10(-6) M bovine PTH-(1-34). This response was inhibited by the PTH analog and antagonist Nle8,18,Tyr34-bPTH-(3-34). We also examined whether human dermal fibroblasts are capable of responding to either a human PTH-like tumor-derived factor or the PTH-like factor contained in human keratinocyte-conditioned medium. Both human humoral hypercalcemia of malignancy-associated tumor extract (2.5 X 10(-10) M) and keratinocyte-conditioned medium (8 X 10(-10) M) stimulated human dermal fibroblast adenylate cyclase. These concentrations are markedly lower than those required for PTH-induced adenylate cyclase stimulation. This activity was also inhibited by the PTH analog. The high prevalence of PTH-responsive adenylate cyclase in dermal fibroblast lines and the apparent potency of tumor-derived and keratinocyte-derived PTH-like factors in dermal fibroblasts suggest that these factors may play a role in normal dermal physiology.

Frequency of various congenital heart diseases in Chinese adults: analysis of 926 consecutive patients over 13 years of age.

A retrospective analysis was performed in 926 consecutive Chinese persons, aged 14 to 68 years, in whom congenital heart disease had been diagnosed and catheterization had been performed over the past 24 years. The incidence of various congenital cardiac malformations was compared with that in other series reported world-wide. About 60% of the 926 patients underwent surgery, and anatomic details of the malformations were examined. Congenital aortic valvular stenosis and coarctation of the aorta were uncommon (2.4% and 1.2%, respectively) among the Chinese adults, and the ethnic differences with Western countries in the incidence of these 2 defects were apparent and real. In contrast, aneurysm of the sinus of Valsalva, either ruptured or nonruptured, was more common (4.3%) and the incidence of outlet muscular type (or type 1) of ventricular septal defect was 4 to 9 times as common as has been reported in the West. Genetic differences rather than environmental factors or dietary habits may explain these special features.