Heterogeneity in hormone responses and patterns of collagen synthesis in cloned dermal fibroblasts.

Fibroblasts cultured from normal human dermis are heterogeneous with respect to growth kinetics, synthetic function, and morphologic features. There are many examples of clonal heterogeneity in apparently homogeneous connective tissue cell populations, and it has been suggested that selection of cell populations with particular phenotypic features is the basis for the development of pathologic connective tissue changes in inflammatory disorders. In these studies we report characterization of the pattern of matrix biosynthesis and responses to hormones in cells cloned from normal human dermis. The results indicate that cloned dermal fibroblasts are heterogeneous with respect to synthesis of collagens as well as their responses to prostaglandin E2 and parathyroid hormone. Selective expansion of clonal populations with unique patterns of matrix synthesis and cell surface receptors could provide the basis for abnormal connective tissue remodeling in certain pathologic states.

Assessing progression from mild to severe preeclampsia in expectantly managed preterm parturients.

OBJECTIVE: To identify factors associated with shortened latency in expectantly managed women diagnosed with preeclampsia without severe features between 23 0/7 and 35 6/7weeks' gestation. METHODS: This was a retrospective cohort study performed at a large community-based hospital between 2009 and 2014, evaluating all mothers between 23 0/7 and 35 6/7weeks' gestation with a diagnosis of preeclampsia without severe features. We collected maternal demographics, symptoms, vital signs and laboratory values within six hours of admission. Maternal and neonatal outcomes were stratified by latency period of less than or greater than/equal to seven days (<7d or ⩾7d). RESULTS: Overall mean latency was 7.6±12.1days. When stratifying subjects to <7d or ⩾7d latency, neither maternal demographics nor gestational age at diagnosis differed between groups. For subjects with ⩾7d latency, pregnancy was prolonged by a mean of 24days compared to the <7d latency group (34 1/7 vs 30 4/7weeks GA, P=0.001). Systolic blood pressure greater than 160mmHg within the first six hours of hospital presentation correlated with a more than 3-fold risk for requiring delivery within seven days of diagnosis (OR 3.26 95% CI 1.40-7.58). CONCLUSION: Within our cohort of preterm women admitted with preeclampsia without severe features, elevated systolic blood pressure on admission conveyed significant risk for delivery within seven days.

A randomized controlled trial of differing doses of postcesarean enoxaparin thromboprophylaxis in obese women.

OBJECTIVE: To compare two enoxaparin dosing strategies at achieving prophylactic anti-Xa levels in women with a body mass index (BMI) ⩾35 (kg m(-2)) postcesarean delivery. STUDY DESIGN: Women with BMI ⩾35 were randomized to receive prophylactic enoxaparin at a fixed dose of 40 mg daily or weight-based dosing of 0.5 mg kg(-1) twice daily. The primary outcome was the proportion of subjects with peak anti-Xa levels in the prophylactic range of 0.2 to 0.6 IU ml(-1). RESULT: From August 2013 through February 2014, 84 demographically similar women completed the protocol. In the weight-based group, 88% (37/42) of the women reached prophylactic anti-Xa levels versus 14% (6/42) in the fixed dose group (odds ratio 44.4, 95% confidence interval 12.44, 158.48, P<0.001). No anti-Xa level exceeded 0.48 IU ml(-1). There were no venous thromboembolic or bleeding events requiring reoperation or transfusion in either group. CONCLUSION: Compared with fixed dosing daily, weight-based dosing twice daily more effectively achieved prophylactic anti-Xa levels without reaching the therapeutic range.

Immune interferon inhibits collagen synthesis by rheumatoid synovial cells associated with decreased levels of the procollagen mRNAs.

Recombinant immune interferon, (interferon-gamma, IFN-gamma) inhibits types I and III collagen synthesis by rheumatoid synovial fibroblast-like cells in culture. This decrease is associated with a decrease in the levels of types I and III procollagen mRNAs in these cells as measured by dot blot hybridization. In the control synovial cells the level of alpha 2(I) mRNA is disproportionately high compared with that of alpha 1(I) or alpha 1(III) mRNA, and IFN-gamma suppresses the level of alpha 1(I) and alpha 1(III) mRNA to a greater extent than that of alpha 2(I) mRNA. The lymphokine, IFN-gamma, may thus have a role in the regulation of collagen synthesis in inflammatory joint disease and other conditions.

Mechanisms of matrix degradation in rheumatoid arthritis.

In the inflammatory synovium production of collagenase is probably responsible for the degradation of collagen in the extracellular matrix and distortion of the architecture and function of the joints. Major collagenase-producing cells are mesenchymal cells such as fibroblasts and chondrocytes, which synthesize and secrete the enzyme influenced by the action of cytokines produced by adjacent mononuclear cells. The cytokines act primarily through cell-surface receptors, whose signal is probably then mediated by complexes of nuclear oncoproteins, to activate transcription of the procollagenase gene. The increased production of collagenase ultimately is the result of a cascade of cellular effects involving complex interactions of different ligands in a system characterized by amplification and feedback loops.

Novel heterozygous ABCB4 gene mutation causing recurrent first-trimester intrahepatic cholestasis of pregnancy.

Intrahepatic cholestasis of pregnancy (ICP) typically presents in the late second or third trimester and carries an increased risk of fetal demise and neonatal morbidity and mortality. First trimester onset is rare and should alert the physician to explore a possible genetic basis for the disease. We present a 26-year-old Hispanic gravida 3, para 0202 with recurrent first-trimester onset ICP. Given her atypical history and presentation, a genetic cause was considered. She was found to have a novel heterozygous missense mutation in the ABCB4 canalicular membrane transport gene. First or early second trimester presentation of ICP should prompt investigation into genetic causes of the disease. Individualized family counseling and neonatal evaluation should be addressed if a disease-causing genetic mutation is diagnosed.

Inhibition of Rous sarcoma virus replication and cell transformation by a specific oligodeoxynucleotide.

The tridecamer d(A-A-T-G-G-T-A-A-A-A-T-G-G), which is complementary to 13 nucleotides of the 3'- and 5'-reiterated terminal sequences of Rous sarcoma virus 35S RNA, was added to chick embryo fibroblast tissue cultures infected with Rous sarcoma virus. Inhibition of virus production resulted. The inference emerges that the tridecamer and its counterpart with blocked 3'- and 5'-hydroxyl termini enter the chick fibroblast cells, hybridize with the terminal reiterated sequences at the 3' and 5' ends of the 35S RNA, and interfere with one or more steps involved in viral production and cell transformation. Likely sites of action are (i) the circularization step of the proviral DNA intermediate, and (ii) the initiation of translation, the latter being described in the following communication [Stephenson, M. L. & Zamecnik, P. C. (1978) Proc. Natl. Acad. Sci. USA 75, 285--288].

Inhibition of Rous sarcoma viral RNA translation by a specific oligodeoxyribonucleotide.

A tridecamer oligodeoxynucleotide, d(A-A-T-G-G-T-A-A-A-A-T-G-G), which is complementary to reiterated 3'- and 5'-terminal nucleotides of Rous sarcoma virus 35S RNA, is an efficient inhibitor of the translation of proteins specified by the viral RNA in the wheat embryo cell-free system. The inhibition specificity for oncornavirus RNA is greater than for rabbit reticulocyte mRNA or brome mosaic virus RNA. Other oligodeoxynucleotides of similar size have little or no specific effect on the RNA-directed translation. The tridecamer acts as a primer for the avian myeloblastosis virus DNA polymerase when Rous sarcoma virus heated 70S RNA is used as a template, offering evidence that it can hybridize to the RNA. The possible use of such an oligodeoxynucleotide hybridization competitor to inhibit Rous sarcoma virus replication is described in the preceding paper [Zamecnik, P. C. & Stephenson, M. L. (1978) Proc. Natl. Acad. Sci. USA. 75, 280--284].

Mechanism of stimulation of prostaglandin synthesis by a factor from rheumatoid synovial tissue.

Rheumatoid synovial cell monolayers, with [1-14C]arachidonic acid ([1-14C]AA) incorporated into cell lipids, are stimulated by a factor (RSF) produced by explant cultures of rheumatoid synovial tissue to produce up to 50-fold increases in [1-14C]prostaglandin E2 and [1-14C]prostaglandin I2. In contrast, levels of free [1-14C]AA released from RSF-stimulated cells are generally lower than [1-14C]AA levels in cultures of untreated cells. These observations are inconsistent with a mechanism of prostaglandin stimulation consisting of an increase in phospholipase activity, because this mechanism would increase free AA levels as well as prostaglandins. A mechanism is proposed in which free AA is maintained at low steady-state levels by reacylation of free AA into phospholipids at a rate more rapid than its reaction with cyclooxygenase to form prostaglandins. In this mechanism, stimulation of the rate of the cyclooxygenase step by RSF accounts for increased prostaglandin synthesis as well as the decreased release of AA. On the basis of data previously reported by others, it is suggested that this mechanism may also be applicable to the stimulation of prostaglandin synthesis by several other agents. Preliminary characterization of the RSF indicates that it is a protein, and molecular seive chromatography indicates that its molecular weight is about 18,000. The production of RSF by rheumatoid synovial tissue is suppressed to undetectable levels by 1 microM dexamethasone.

Immune interferon suppresses levels of procollagen mRNA and type II collagen synthesis in cultured human articular and costal chondrocytes.

Cultured human articular and costal chondrocytes were used as a model system to examine the effects of recombinant gamma-interferon (IFN-gamma) on synthesis of procollagens, the steady state levels of types I and II procollagen mRNAs, and the expression of major histocompatibility complex class II (Ia-like) antigens on the cell surface. Adult articular chondrocytes synthesized mainly type II collagen during weeks 1-3 of primary culture, whereas types I and III collagens were also produced after longer incubation and predominated after the first subculture. Juvenile costal chondrocytes synthesized no detectable alpha 2(I) collagen chains until after week 1 of primary culture; type II collagen was the predominant species even after weeks of culture. The relative amounts of types I and II collagens synthesized were reflected in the levels of alpha 1(I), alpha 2(I), and alpha 1(II) procollagen mRNAs. In articular chondrocytes, the levels of alpha 1(I) procollagen mRNA were disproportionately low (alpha 1(I)/alpha 2(I) less than 1.0) compared with costal chondrocytes (alpha 1 (I)/alpha 2(I) approximately 2). Recombinant IFN-gamma (0.1-100 units/ml) inhibited synthesis of type II as well as types I and III collagens associated with suppression of the levels of alpha 1(I), alpha 2(I), and alpha 1(II) procollagen mRNAs. IFN-gamma suppressed the levels of alpha 1(I) and alpha 1(II) procollagen mRNAs to a greater extent than alpha 2(I) procollagen mRNA in articular but not in costal chondrocytes. Human leukocyte interferon (IFN-alpha) at 1000 units/ml suppressed collagen synthesis and procollagen mRNA levels to a similar extent as IFN-gamma at 1.0 unit/ml. In addition, IFN-gamma but not IFN-alpha induced the expression of HLA-DR antigens on intact cells. The lymphokine IFN-gamma could, therefore, have a role in suppressing cartilage matrix synthesis in vivo under conditions in which the chondrocytes are in proximity to T lymphocytes and their products.

Ventilation conditions and air-borne bacteria and particles in operating theatres: proposed safe economies.

Concentrations of air-borne bacteria and particles have been measured in turbulently ventilated operating theatres in full flow, half flow and zero flow conditions. Increased air-borne challenge produced by human activity and by mechanical cleaning procedures is demonstrated: die-away of this contamination is shown to be related to the ventilation rate. Ventilation can be reduced or turned off at night and during weekends, and cleaning can also be carried out, without increased risk of infection if full flow is restored one hour prior to preparation for surgery. Areas surrounding the theatres should remain at positive pressure with regard to the general hospital environment during low or no flow periods. The implementation of such energy-saving policies will substantially reduce theatre running costs without introducing infection hazards.

The 3'-terminal nucleosides of the high molecular weight RNA of avian myeloblastosis virus.

The RNA isolated from avian myeloblastosis virus was fractionated by sucrose density gradient centrifugation. The 3'-OH terminal nucleosides of various fractions were determined by periodate oxidation followed by tritiated borohydride reduction. The 60-70S fraction and the 35S RNA derived from it by heating both have adenosine as the major terminal nucleoside, with cytidine as the next most frequent terminal. Control samples of tRNA(met) (f. coli) and 28S ribosomal RNA from mouse ascites tumor cells gave the expected terminal residues and molecular weights.

Is screening for bacteriuria in pregnancy worth while?

A total of 4470 pregnant women were screened for bacteriuria by the dipslide method and significant growth found in 226 (5.1%). In 198 cases the urine was re-examined, in 119 by using suprapubic aspiration or catheterisation (62 (52%) samples contained bacteria) and in 79 by using midstream urine samples (26 (33%) samples contained greater than 10(8) colony forming units/1), showing the maximum prevalence of confirmed bacteriuria to be 2.6%. Overt urinary tract infection developed later in four of 80 patients with proved bacteriuria who had been given antibiotics, in one of eight untreated patients with bacteriuria, in one of 110 patients with unconfirmed bacteriuria, and in one of 226 non-bacteriuric controls. A history of urinary tract infection was given by 18% of controls and 42% of women with confirmed bacteriuria. Screening for bacteriuria and treatment with antibiotics to prevent later overt infection is expensive. Whether it is worth while and cost effective depends largely on the prevalence of bacteriuria in the local population and the proportion who develop overt infection. The screening and treatment programme reported here appeared to prevent only six cases of overt infection.

Campylobacter pyloridis in peptic ulcer disease: microbiology, pathology, and scanning electron microscopy.

After the recent successful isolation of spiral organisms from the stomach this paper presents the bacteriological and pathological correlation of gastric antral biopsies from 51 patients endoscopied for upper gastrointestinal symptoms. Campylobacter pyloridis was cultured from 29 patients and seen by either silver staining of the biopsy or scanning electron microscopy in an additional three. The organism was cultured from 23 of the 33 (69%) patients with peptic ulcer disease and from within this group 17 (80%) of the 21 patients with duodenal ulceration. It was cultured only once from the 12 normal biopsies in the series but from 27 of the 38 (71%) biopsies showing gastritis. C pyloridis was also cultured from five out of seven of the 14 endoscopically normal patients, who despite this had biopsy evidence of gastritis. It was the sole organism cultured from 65% of the positive biopsies and scanning electron microscopy invariably revealed it deep to the surface mucus layer. C pyloridis persisted in the three patients with duodenal ulcers after treatment and healing. The findings support the hypothesis that C pyloridis is aetiologically related to gastritis and peptic ulceration though its precise role still remains to be defined.

Analysis of oncornavirus RNA subunits by electron microscopy.

Subunits of oncornavirus (avian myeloblastosis virus) RNA were isolated from purified 60--70S viral RNA by heat dissociation. Molecules sedimenting at 35 S, assumed to be the major component of the viral genome, were visualized in the electron microscope and their lengths were statistically analyzed. The results indicate a rather heterogeneous population of molecules with five distinct, reproducible size groups, an observation that excludes the assumption of random degradation of the genome. In addition, molecules of 28 and 18S RNA, always present in oncornavirus RNA preparations, were examined with the same methods. Some of these molecules possess secondary-structure regions similar to those characteristic for ribosomal RNA.

Stimulation of procollagenase synthesis parallels increases in cellular procollagenase mRNA in human articular chondrocytes exposed to recombinant interleukin 1 beta or phorbol ester.

Interleukin 1, a product predominantly of monocytes, increases the synthesis and release of procollagenase and prostaglandin E2 by mesenchymal target cells such as synovial fibroblasts and articular chondrocytes, an effect mimicked by some phorbol esters. In order to determine the mechanisms underlying these responses primary cultures of human articular chondrocytes were preincubated with recombinant human interleukin 1 beta or the phorbol ester, phorbol 12-myristate 13-acetate, in the presence or absence of the cyclooxygenase inhibitor, indomethacin. Interleukin 1 beta or phorbol ester increased the levels of procollagenase (assayed after trypsin activation) and the labeling of several medium proteins by cells incubated with [35S]methionine, independent of prostaglandin synthesis. The labeling of a 55 kD protein immunocomplexed with antibodies to procollagenase was also increased. The increased synthesis of procollagenase was paralleled by increased cellular levels of procollagenase mRNA, determined with a cDNA probe coding for human procollagenase. Thus the increased synthesis of procollagenase in response to the inflammatory mediator, interleukin 1, is controlled at a pretranslational level, possibly at the level of transcription.