Identification of diagnostic biomarkers and immuno-infiltration analysis for rheumatoid arthritis based on biological information and WGCNA.

OBJECTIVE: Rheumatoid arthritis (RA), as an autoimmune disease, poses a huge social and economic burden worldwide. Although the diagnosis of RA has been gradually improved, there is still a need to discover accurate and rapid biomarkers for diagnosis and therapy with a precise understanding of the disease. This study aimed to screen diagnostic biomarkers and analyze immune infiltration in RA based on weighted gene co-expression network analysis (WGCNA). MATERIALS AND METHODS: Firstly, we screened the experimental and validation sets associated with RA from the GEO database. Crossover genes were obtained using differential genes (DEGs) and key modules in WGCNA. Subsequently, the crossover genes were constructed into protein-protein interaction (PPI) networks and screened to obtain hub genes. The receiver operating characteristic (ROC) curve assessment was performed to identify diagnostic biomarkers. In addition, we used the Cibersort algorithm for immuno-infiltration analysis and the DGidb database to search for drugs associated with diagnostic biomarkers. RESULTS: In the end, 377 DEGs were identified, and the enrichment analysis revealed significant associations with the immune system. Blue modules in the WGCNA analysis were positively associated with the disease and were identified as key modules. ROC curves evaluated the four hub genes, which significantly differentiated RA from healthy controls and could be used as diagnostic biomarkers. In further analysis, we found that RA is closely related to immunity, and the search identified multiple drugs that hold promise for treating RA. CONCLUSIONS: BCL2A1, PTGS2, FAS, and LY96 may be used as diagnostic biomarkers, which is significant for diagnosing and treating RA.

[The citation analysis of the articles published in Chinese Journal of Otorhinolaryngology Head and Neck Surgery from 2009 to 2019].

Objective: To analyze the citation of articles in Chinese Journal of Otorhinolaryngology Head and Neck Surgery, and to explore the strategy for running scientific journals. Methods: Using the Citation Database of Chinese Biomedical Journals, the citations of articles in this journal from 2009 to 2019 were statistically analyzed, and the characteristics of highly cited articles were analyzed. Results: From 2009 to 2019, Chinese Journal of Otorhinolaryngology Head and Neck Surgery published 2 814 papers. Cited papers accounted for 75.69% of all papers, and each paper was cited 7.24 times. The citation rate of published papers was consistent with the variation trend of citation frequency. Non treatises such as guide consensus and original articles for special issues were cited better. There was no obvious correlation between paper funding and citation. Conclusion: Improving the content quality and optimizing the column setting are important measures to enhance the influence of scientific journals.

Health behaviour practices and expectations for a local cancer survivorship programme: a cross-sectional study of survivors of childhood cancer in Hong Kong.

INTRODUCTION: Lifestyle choices may influence health outcomes in cancer survivors. This study of childhood cancer survivors in Hong Kong investigated factors associated with health-protective and health-damaging behaviours; it also examined expectations of a survivorship programme. METHODS: This cross-sectional study recruited survivors of childhood cancer ≥2 years after treatment. Survivors completed a structured questionnaire to report their health practices and the perceived values of survivorship programme components. Multivariable logistic regression analysis was conducted to identify factors associated with health behaviours. RESULTS: Two hundred survivors were recruited (mean age=23.4 ± 8.8 years; mean duration since treatment, 13.4 ± 7.6 years). Comparatively few survivors exercised ≥4 days/week (16.0%), used sun protection (18.0%), and had a balanced diet (38.5%). Furthermore, comparatively few survivors reported that they had not undergone any immunisation (24.5%) or were unsure (18.5%) about their immunisation history. Most adult survivors were never-drinkers (71.0%) and never-smokers (93.0%). Brain tumour survivors were more likely to have unhealthy eating habits, compared with haematological malignancy survivors (odds ratio [OR]=2.45; 95% confidence interval [CI]=1.29-4.68). Lower socioeconomic status was associated with inadequate sun protection (OR=0.20; 95% CI=0.05-0.83), smoking (OR=5.13; 95% CI=1.48-17.75), and exposure to second-hand smoke (OR=3.52; 95% CI=1.42-8.69). Late-effects screening (78.5%) and psychosocial services to address psychological distress (77%) were considered essential components of a survivorship programme. CONCLUSIONS: Despite the low prevalences of health-damaging behaviours, local survivors of childhood cancer are not engaging in health-protective behaviours. A multidisciplinary programme addressing late effects and psychosocial aspects may address the multifaceted needs of this special population.

Outcome of behavioural treatment for idiopathic chronic constipation.

BACKGROUND/AIM: Behavioural therapy is effective in patients with chronic intractable constipation despite standard treatment, but long-term results in unselected patients are unclear. This study investigates the effects of behavioural therapy on symptoms, subjective well-being, and the physical and mental quality of life. METHODS: Patients who had failed standard care for idiopathic chronic constipation underwent behavioural therapy in a specialist clinic. Symptom severity and quality of life were assessed before and after therapy using the 'Constipation Scoring System' and the Short-Form 36 questionnaire. The primary outcome was subjective perception of improvement. Secondary outcomes were symptoms of constipation and quality of life scores. RESULTS: Of 233 consecutive patients with self-reported constipation (median symptom duration 5-10 years, median age 44 years, females 86%), 180 (77%) completed treatment in a median of three (range 1-7) sessions. One hundred and sixty-five patients (71% of all referrals or 92% of those completing treatment) reported subjective improvement. Median bowel frequency improved from once every 2-7 days to 1-3 per day (P = 0.05). Pain and bloating improved in more than 80% of patients. The Short-Form 36 physical (P < 0.05) and mental (P < 0.05) composite scores improved significantly. Patients with a longer duration of symptoms were less likely to complete treatment. Digital evacuation prior to treatment was a predictor of poor outcome. CONCLUSION: Behavioural therapy is associated with significant improvement in symptoms of chronic constipation and quality of life. Non-drug therapies that successfully treat patients with functional gut disorders resistant to standard treatment are needed in the mainstream provision of care.

A comprehensive system for protein purification and biochemical analysis based on antibodies to c-myc peptide.

The genomics revolution has created a need for increased speed and generality for recombinant protein production systems as well as general methods for conducting biochemical assays with the purified protein products. 9E10 is a well-known high-affinity antibody that has found use in a wide variety of biochemical assays. Here we present a standardized system for purifying proteins with a simple epitope tag based on c-myc peptide using an antibody affinity column. Antibodies with binding parameters suitable for protein purification have been generated and characterized. To purify these antibodies from serum-containing medium without carrying through contaminating immunoglobulin G, a peptide-based purification process was developed. A fluorescence polarization binding assay was developed to characterize the antigen-antibody interaction. Protein purification protocols were optimized using a fluorescein-labeled peptide as a surrogate "protein." Binding and elution parameters were evaluated and optimized and basic operating conditions were defined. Several examples using this procedure for the purification of recombinant proteins are presented demonstrating the generality of the system. In all cases tested, highly pure final products are obtained in good yields. The combination of the antibodies described here and 9E10 allow for almost any biochemical application to be utilized with a single simple peptide tag.

Microsatellite alterations on human chromosome 11 in in situ and invasive breast cancer: a microdissection microsatellite analysis and correlation with p53, ER (estrogen receptor), and PR (progesterone receptor) protein immunoreactivity.

BACKGROUND AND OBJECTIVES: Microsatellite instability (MSI) has been documented in a subset of sporadic tumors. Loss of heterozygosity (LOH) on chromosome 11 loci in breast cancer is a frequent event. The purpose of the present study is to examine the incidence of microsatellite alterations in in situ and invasive human breast carcinoma and to clarify their significance in regulating the dynamics of cancer progression. METHODS: Four highly polymorphic (CA)n repeat microsatellites were used to determine microsatellite alterations in ten ductal carcinoma in situ (DCIS) and 19 invasive ductal carcinoma (IDC). To investigate the expression of p53, ER (estrogen receptor), and PR (progesterone receptor) association with MSI, immunohistochemistry staining was applied. RESULTS: MSI were detected in 20% (2/10) of DCIS and in 47.4% (9/19) of IDC. The frequency of MSI in IDC was significantly higher than that in DCIS (P < 0.001). Also, the MSI seemed to correlate with clinical stage (P = 0.0001) and tumor size (P = 0.004) but not histological grade or age. In addition, we found that 27% of the tumors showed LOH at 11q23.3-24 region between loci D11S934 and D11S912. Seven of nine MSI cases demonstrated low or no expression of p53. However, there was significantly reduced expression of PR, but not ER in MSI cases. CONCLUSIONS: Our results suggest that breast cancer acquires the RER phenotype (replication-error phenotype) in the relatively late stages, and that the RER phenotype is associated with aggressiveness of IDC (infiltrative duct carcinoma). The result also implicated that mismatch repair failure can alter the expression of PR but not ER and p53.

[A study on thermal decomposition mechanism and quality analysis of pearl powder].

OBJECTIVE: To establish a rapid and convenient method for distinguishing genuine from sham of pearl powder as well as appraising its quality preliminarily. METHOD: Thermogravimetry and differential thermogravimetry. RESULT: The TG and DTG curves can be divided into two characteristic regions. The first step ranges from 250 to 380 degrees C with a weight loss of about 3%, resulting from the denaturalization and decomposition of organic matter in the powder; and the second step from 600 to 780 degrees C with a weight loss of about 40% resulting from the decomposition of calcium carbonate in the powder. CONCLUSION: According to the two characteristic regions on TG and DTG curves along with corresponding parameters, pearl powder can be appropriately authenticated. Being related directly to the contents of organic matter in pearl powder, the first step is an important criterion for quality appraisal.

Longitudinal analysis of methicillin-resistant Staphylococcus aureus isolates at a teaching hospital in Taiwan.

In Taiwan, the frequency of nosocomial infections caused by methicillin-resistant Staphylococcus aureus (MRSA) has increased rapidly during the past 10 years. To investigate the epidemiology of MRSA infections, a total of 140 MRSA isolates collected at National Taiwan University Hospital from 1992 to 1996 were characterized by pulsed-field gel electrophoresis (PFGE) profiles and antibiotypes, as determined with the disk diffusion method. Among these isolates, six PFGE types (with 20 subtypes) and six antibiotypes were identified. Antibiotyping proved to be a poor method of epidemiologic analysis, because almost all of the MRSA isolates analyzed shared a very similar multidrug-resistant antibiotype. Most MRSA infections and colonizations in this hospital were due to the spread of strains belonging to three major PFGE types (A, B, and C). However, the major type changed in different years with types A, B, and C being predominant in 1992 through 1993, 1994 through 1995, and 1996, respectively. The three major PFGE types spread easily throughout the hospital wards, presumably carried by health care workers and environmental contamination. Our results demonstrate that there was a dominant strain spreading in our hospital each year and the dominant strain may shift in different years.

Ca2+ and Mg2+ selectively induce aggregates of PHF-tau but not normal human tau.

The molecular mechanism of pathological aggregation of microtubule-associated protein tau during neurodegeneration is unclear. In the present study, the in vitro effect of various metal ions on the aggregation of tau was examined using paired helical filament tau (PHF-tau) obtained from corticobasal degeneration (CBD) and Alzheimer's disease (AD) brains as well as normal human tau proteins isolated from fetal and adult brains and a recombinant system. Among the metal ions tested, Ca2+ and Mg2+ effectively induced formation of approximately 340 kD aggregates of PHF-tau but not normal tau proteins as determined by sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis and immunoblotting. Al3+ and Fe2+ precipitated both PHF-tau and normal tau protein as SDS-insoluble pellets. The other metal ions examined (Cu2+, Zn2+, and Li+) were inactive and caused neither aggregation nor precipitation of any tau protein. Intermixing experiments using PHF-tau and various normal tau preparations showed that the 340-kD aggregates induced by Ca2+ contained PHF-tau but not normal tau regardless whether unmodified (recombinant) or highly phosphorylated (fetal brain) tau proteins were used. The present results suggest that post-translational modifications other than the fetal-type phosphorylation are required for Ca2+- and Mg2+-dependent aggregation of PHF-tau and that the regional elevation of these ions may trigger pathological deposition of PHF-tau in certain neurodegenerative disorders.

Hyaluronate binding assay study of transfected CD44 V4-V7 isoforms into the human gastric carcinoma cell line SC-M1.

The potential human metastasis molecule CD44 and its isoforms V5 and V6 are overexpressed in human gastric carcinoma. Among the numerous extracellular matrix components, hyaluronate, a CD44 ligand, is of increasing interest in relation to its role in cancer cell development and invasion. By using the dynabead separation method, the SC-M1 cell line was separated into V5 and V6 isoform-positive and -negative populations. The V5 and V6 isoform-negative populations exhibited significantly higher hyaluronate binding activity than the corresponding positive cells. The hyaluronate binding activity of V5 and V6-positive cells could be restored by pretreatment with anti-CD44 V5 and V6 monoclonal antibodies (MAbs). In addition, transfection of aVV5 and V6-negative cells decreased their hyaluronate binding activity to the levels of CD44 V5 and V6-positive cells. Cells transfected with V5 and V6 recovered their hyaluronate binding activity after pretreatment with MAbs against V5 and V6. These data suggest that cell adhesion involving hyaluronate can be regulated by multiple mechanisms, one of which involves alternative splicing of CD44 isoforms.

Tau released from paired helical filaments with formic acid or guanidine is susceptible to calpain-mediated proteolysis.

Paired helical filaments (PHFs), a characteristic neuropathologic finding in Alzheimer's disease brain, are abnormal fibrillary forms of hyperphosphorylated tau (PHF-tau), which have been shown to be highly resistant to calpain digestion. Either excessive phosphorylation or fibrillary arrangement of tau proteins in PHFs may play a role in proteolytic resistance by limiting access to calpain recognition/digestion sites. To determine the contribution of the fibrillary conformation, isolated PHFs were subjected to treatment with either formic acid or guanidine. Both procedures effectively abolished the fibrillary structure of PHF but preserved PHF-tau immunoreactivity using a panel of antibodies that recognize nonphosphorylated and phosphorylated epitopes. These treatments also significantly increased the sensitivity of PHF-tau polypeptides to calpain proteolysis as shown by significant decreases in the half-life (t(1/2)) from the infinite with native PHF to 44 min and 4.4 min in formic acid- or guanidine-treated samples, respectively. In contrast, the sensitivity of normal fetal tau (3.4 min) was either decreased (5.9 min) or unaffected (3.6 min) by similar treatment. Our results indicate that after guanidine treatment, the sensitivity of PHF to calpain resembles that of fetal tau. These results strongly suggest that the fibrillary structure of PHF-tau, rather than hyperphosphorylation, is the major factor responsible for the resistance of abnormal filaments to calpain-mediated proteolysis.

Secular trends in the epidemiology of nosocomial fungal infections at a teaching hospital in Taiwan, 1981 to 1993.

OBJECTIVE: To describe the incidence and patterns of nosocomial fungal infection in a large teaching hospital in Taiwan. DESIGN: Prospective, hospitalwide nosocomial surveillance data from 1981 through 1993 were analyzed to show the secular trend in nosocomial fungal infection rates and to identify the most common pathogens and sites of infection (other than skin) in this hospital. SETTING AND PATIENTS: The National Taiwan University Hospital is a medical school-affiliated hospital in the city of Taipei, Taiwan, with a 1200-bed capacity before 1991 and 1500 beds since 1992. It provides both primary and tertiary medical care. RESULTS: The overall nosocomial fungal infection rate rose from 0.9 infections per 1000 discharges in 1981 to 6.6 per 1000 discharges in 1993, with the highest rate at the medical intensive-care unit (26.5/1000 discharges in 1993). This increase in infection rate was found at four major anatomic sites of infection, particularly including the bloodstream (0.08-2.19/1000 discharges) and the urinary tract (0.36-2.95/1000 discharges). Of 256 pathogens causing nosocomial fungemia from 1981 through 1993, Candida albicans was the most commonly isolated (50.8%), followed by Candida tropicalis (17.6%). Candida parapsilosis (11.7%), and Candida glabrata (8.2%). As compared to isolates from 1981 through 1988, the proportion of C parapsilosis and C glabrata isolated between 1989 and 1993 increased more than sixfold and fourfold, respectively. The increasing importance of fungal infections was confirmed further by the increased use of amphotericin B and azoles in this hospital. CONCLUSIONS: Candida species and other yeasts have become a prominent cause of nosocomial infections in this hospital. These fungal pathogens accounted for a higher proportion of nosocomial bloodstream and urinary infections than any single bacterial species. Therefore, it is important to conduct a prospective epidemiological study and to establish in vitro antifungal susceptibility testing to enhance efforts to control nosocomial fungal infections and to minimize the risk of emergence of antifungal resistance.

Increasing nosocomial infections of methicillin-resistant Staphylococcus aureus at a teaching hospital in Taiwan.

Methicillin-resistant Staphylococcus aureus (MRSA) is a worldwide important pathogen in nosocomial infections. To investigate the extent of the problem in Taiwan, analysis for the period of 1981-1994 was carried out of prospective surveillance data from the National Taiwan University Hospital, a major university teaching hospital in Taiwan. The number of nosocomial MRSA infections increased from five in 1981 to 133 in 1994, and the incidence increased from 0.2 episodes/1000 discharges in 1981 to 2.9 episodes/1000 discharges in 1994. The most common infection site was surgical wounds, which accounted for 26.3% of total 577 episodes of nosocomial MRSA infections during the study period. However, bacteraemia has become more and more common during the past 14 years. MRSA infections occured more frequently in patients stayed in the burn unit and other intensive care units than in the general wards. Other than oxacillin, the resistance rate to many other antibiotics also increased in S. aureus strains causing nosocomial infections in this hospital. Vancomycin remained active to all these S. aureus strains, even until 1994.

Ultrastructural instability of paired helical filaments from corticobasal degeneration as examined by scanning transmission electron microscopy.

Paired helical filaments (PHFs) accumulate in the brains of subjects affected with Alzheimer's disease (AD) and certain other neurodegenerative disorders, including corticobasal degeneration (CBD). Electron microscope studies have shown that PHFs from CBD differ from those of AD by being wider and having a longer periodicity of the helical twist. Moreover, PHFs from CBD have been shown to be primarily composed of two rather than three highly phosphorylated polypeptides of tau (PHF-tau), with these polypeptides expressing no exons 3 and 10. To further explore the relationship between the heterogeneity of PHF-tau and the appearance of abnormal filaments, the ultrastructure and physical parameters such as mass per unit length and dimensions were compared in filaments from CBD and AD using high resolution scanning transmission electron microscopy (STEM). Filament-enriched fractions were isolated as Sarcosyl-insoluble pellets and for STEM studies, samples were freeze-dried without prior fixation or staining. Ultrastructurally, PHFs from CBD were shown to be a heterogeneous population as double- and single-stranded filaments could be identified based on their width and physical mass per unit length expressed in kilodaltons (kd) per nanometer (nm). Less abundant, double-stranded filaments had a maximal width of 29 nm and a mass per unit length of 133 kd/nm, whereas three times more abundant single-stranded filaments were 15 nm wide and bad a mass per unit length of 62 kd/nm. Double-stranded filaments also displayed a distinct axial region of less dense mass, which appeared to divide the PHFs into two protofilament-like strands. Furthermore, these filaments were frequently observed to physically separate along the long axis into two single strands or to break longitudinally. In contrast, PHFs from AD were ultrastructurally stable and uniform both in their width (22 nm) and physical mass per unit length (104 kd/nm). The ultrastructural features indicate that filaments of CBD and AD differ both in stability and packing of tau and that CBD filaments, composed of two distinct protofilaments, are more labile under STEM conditions. As fixed and stained filaments from CBD have been shown to be stable and uniform in size by conventional transmission electron microscopy, STEM studies may be particularly suitable for detecting instability of unstained and unfixed filaments. The results also suggest that molecular heterogeneity and/or post-translational modifications of tau may strongly influence the morphology and stability of abnormal filaments.

Calpain-induced proteolysis of normal human tau and tau associated with paired helical filaments.

The major components of neurofibrillary tangles (NFT) in Alzheimer's disease are bundles of paired helical filaments (PHF) which are primarily composed of highly phosphorylated tau proteins (PHF-tau). To further understand the mechanism of PHF accumulation in NFT, we examined the calpain-induced proteolysis of highly purified and primarily non-aggregated PHF and normal tau proteins with various contents of phosphate isolated from either fetal (F-tau) or adult human brain (N-tau). The extent of proteolysis was determined by decreases in tau immunoreactivity using Western-blot analysis and a panel of site-specific tau antibodies (Alz 50, Tau-2, Tau 14, Tau-1, AT8, E-11, AH-1 and PHF-1). We found that full-size polypeptides of N-tau and F-tau were similarly and rapidly proteolyzed in vitro by calpain (calpain II, 3.3 units/mg protein) during a 10-min incubation at 30 degrees C, and that their half lives (t1/2) were 1.5 min and 1.8 min, respectively. Analysis of immunoblots suggests that full-length polypeptides of tau are first degraded into large fragments similar in size to that generated endogenously, then into smaller fragments. Since both endogenous and in-vitro-generated tau fragments retained N-terminal epitopes, the results suggest that most of the calpain-sensitive sites may be located in the C-terminal half of the tau molecule. In contrast, PHF were extremely resistant to degradation and only a fivefold higher concentration of calpain (16.7 units/mg protein) induced partial proteolysis of PHF. A major calpain-generated fragment was a 45-kDa polypeptide derived from the C-terminal region of PHF-tau, which forms a core of filaments. The results suggest that the inaccessibility of potential calpain-digestion sites in the filament core could contribute to the resistance of PHF to calpain and subsequently lead to the accumulation of PHF in Alzheimer's disease. The results also suggest that hyperphosphorylation of tau may be marginally involved in the resistance of PHF to degradation by calpain. Ultrastructural examination revealed that, in contrast to previous studies with trypsin, calpain did not alter the morphologic appearance of filaments; after incubation with calpain, the majority of PHF remained short and disperse and the number of PHF aggregated into NFT-like clusters was not significantly increased. The results suggest that the role of calpain in promoting the aggregation and clustering of filaments is limited.

[Secular trends in the etiology of nosocomial infection at a teaching hospital in Taiwan, 1981-1994].

Surveillance system of nosocomial infection was established in 1980 at the National Taiwan University Hospital (NTUH). To identify pathogens and the secular trends in the etiology of nosocomial infection from 1981 to 1994, the prospective, hospital-wide nosocomial surveillance data were analysed. During this period, 22,146 pathogens causing nosocomial infections were isolated. Gram-negative aerobic bacteria remained the major pathogens, but gram-positive cocci and fungi increased rapidly in the past 14 years. When the overall pathogen distribution is examined, Pseudomonas areuginosa was the most frequently isolated pathogen, but Candida albicans and other yeasts have taken the leading position since 1993. Staphylococcus aureus and coagulase-negative staphylococci also increase significantly in recent years. When the pathogens causing infection at the 4 major sites were examined. P. aeruginosa was the pathogen most often associated with respiratory tract and surgical wound infections. In blood stream and urinary tract infections, we observed Escherichia coli was replaced by C. albicans and other yeasts as a most common isolate in these years. In addition, C. albicans and other yeasts and methicillin-resistant S. aureus (MRSA) are emerging as major nosocomial pathogens at NTUH. C. albicans and other yeast increased from 1.8% in 1981 to 14.9% in 1994 in the overall nosocomial infection. The increase was found in the blood stream (2.1% to 16.2%) and urinary tract infections (5.4% to 24.7%). Of 1,742 nosocomial S. aureus isolates, the percentage of MRSA rose from 12.5% in 1981 to 55.2% in 1994. The high percentage of MRSA was observed at 4 major anatomic sites of infection. In summary, significant shifts in the pathogens of nosocomial infection have occurred in the past 14 years at NTUH, and the distribution of nosocomial pathogens was similar to those reported in the United States in recent years.

Differential effect of estrogen on the production of cyclin B1, cdc2 p34 and c-fos protein in rat uterus.

The uterine content of c-fos protein, cyclin B1 (cell cycle protein) and cdc2 p34(cyclin-dependent kinase) in immature and mature rats was determined using the enhanced chemiluminescence(ECL) western blot method. Cyclin B1 was found predominantly in immature rat uterus and cdc2 p34 only in mature rat uterus. Several isoforms of c-fos oncogene protein were present in both mature and immature rat uteri. An additional immunoreactive c-fos protein with an estimated molecular weight of 28 kDa was found in mature rat uterus and was missing in immature uterus. Uteri from ovariectomized rats treated with estrogen and/or ICI 182,780, an antiestrogen, were analyzed by ECL western blot. cdc2 p34 and the c-fos 28 kDa protein were found in estradiol-treated rat uteri and were not detected in uteri of control and ICI 182,780-treated animals; whereas Cyclin B1 was absent in uteri from control and estradiol-treated ovariectomized animals. ICI 182,780 administered to estradiol-treated ovariectomized rats blocked the induction of cdc2 p34 and the c-fos 28 kDa protein in the uterus. The present results show that the production of the cell cycle factors, cyclin B1, cdc2 p34 and c-fos, during rat uterine growth are under different regulatory controls. cdc2 p34 and c-fos 28 kDa protein are under the control of estradiol; whereas cyclin B1 and the majority of the immunoreactive isoforms of c-fos are not influenced by this hormone.

Synthesis and secretion of an estrus stage-specific protein by rat uterus.

A specific protein with an estimated molecular weight of 260 kDa was found to be synthesized and secreted into the incubation medium by rat uterus only during the estrus stage of the cycle. This secreted uterine protein was designated as estrus stage-specific protein (ESP). ESP was not produced by pregnant, lactating or immature pup rat uteri. Estradiol administered to ovariectomized rats induced production of ESP which was blocked by the antiestrogen, ICI 182, 780. The present results show that the synthesis and secretion of ESP is regulated by estradiol and this protein maybe involved in blastocyst implantation.

Identification of protein A-binding components in Spisula oocytes.

Components involved in sustaining meiosis arrest of oocytes were determined. Proteins that bind to protein A from meiosis-arrested and 5-HT-matured Spisula oocytes were analyzed by sodium dodecyl sulfate polyacrylamide gel electrophoresis. Meiosis-arrested oocytes contained three doublets of proteins with estimated Mrs of 43 and 45, 38 and 40, and 21 and 23 kDa. In 5 HT-matured oocytes the 21 and 23 and 38 and 40 kDa proteins were retained; whereas the 43 and 45 kDa proteins were absent. The protein A-bound proteins did not interact with antibodies against the various subclasses of human, mouse, rat and rabbit IgG or human Fc fragment. The amino acid sequence of the N-terminus of the 43 kDa protein was determined to be NH2-VLRIGSGMXDT. Comparison of this sequence with existing database at Protein Identification Resource (R 32.0), GenBank (R 72.0), SWISS-PROT (R 22.0), and EMBL (R 31.0) showed no homology with any reported protein. The protein A-bound components from meiosis-arrested oocytes were incubated in vitro with [gamma-32P]ATP. Only the 68 kDa protein was radiophosphorylated. This protein was not detected in 5-HT-matured oocytes. The disappearance of the 43, 45, and 68 kDa proteins in 5-HT-matured oocytes suggests that these components may be involved in sustaining meiosis meiosis. A unique property of these proteins is that they interact with protein A and are distinctly different from immunoglobulin.

Identification of a cAMP-dependent protein kinase in bovine and human follicular fluids.

A soluble protein kinase (PK) was purified from bovine and human follicular fluids (FF) by ultrafiltration through a PM-10 membrane followed by chromatography on heparin-agarose, DEAE-cellulose and cellulose phosphate columns. The PK phosphorylated calf thymus histones and endogenous FF proteins having estimated Mrs of 40, 62, 128 and 180 KD. cAMP enhanced PK activity; whereas protein kinase A (PKA)-inhibitor peptide blocked the activity. The present findings suggest that the enzyme is a cAMP-dependent PK.