A new, nontumorigenic human breast epithelial cell line, HMT-3522, has been established from fibrocystic breast tissue. Cells were explanted and propagated in chemically defined medium including insulin, transferrin, epidermal growth factor, hydrocortisone, estradiol, prolactin, and Na-selenite. The epithelial nature of the cell line was established by immunocytochemical detection of cytokeratins. Moreover, electronmicroscopy revealed monolayers of polarized cells connected by desmosomes and provided with apical microvilli. Milk fat globule membrane antigen, specific for the apical membrane domain of normal, luminal breast epithelial cells, was expressed only in confluent cultures where some cells overlaid others, indicating "stem cell"-like properties. After 25 to 30 passages, the cells are diploid with a few marker chromosomes and loss of chromosomes in the D-group. The cells are nontumorigenic in athymic mice; they lack estrogen receptors, and estradiol does not stimulate growth. The HMT-3522 cell line may represent a useful model for the study of breast cell differentiation and carcinogenesis in vitro.

BACKGROUND

Simple criteria are needed to predict which patients with severe ulcerative colitis will respond poorly to intensive medical treatment and require colectomy.

OBJECTIVE

To find out if the early pattern of change in inflammatory markers or other variables could predict the need for surgery and to evaluate the outcome of medical treatment during one year follow up.

METHODS

51 consecutive episodes of severe colitis (Truelove and Witts criteria) affecting 49 patients admitted to John Radcliffe Hospital, Oxford.

METHODS

Prospective study monitoring 36 clinical, laboratory, and radiographic variables. All episodes treated with intravenous and rectal hydrocortisone and 14 of 51 with cyclosporine.

RESULTS

Complete response in 21 episodes (< or = 3 stools on day 7, without visible blood), incomplete response in 15 >> 3 stools or visible blood on day 7, but no colectomy), and colectomy on that admission in 15. During the first five days, stool frequency and C reactive protein (CRP) distinguished between outcomes (p < 0.00625, corrected for multiple comparisons) irrespective of whether patients or the number of episodes were analysed. It could be predicted on day 3, that 85% of patients with more than eight stools on that day, or a stool frequency between three and eight together with a CRP>> 45 mg/l, would require colectomy. For patients given cyclosporine, four of 14 avoided colectomy but two continued to have symptoms. After admission, complete responders remained in remission for a median nine months and had a 5% chance of colectomy. Incomplete responders had a 60% chance of continuous symptoms and 40% chance of colectomy.

CONCLUSIONS

After three days intensive treatment, patients with frequent stools >> 8/day), or raised CRP >> 45 mg/l) need to be identified, as most will require colectomy on that admission. The role of cyclosporine for treating severe colitis has yet to be defined. After seven days' treatment, patients with>> 3 stools/day of visible blood have a 60% chance of continuous symptoms and 40% chance of colectomy in the following months.

Several properties of lymphoid dendritic cells in situ have been determined, and contrasted to information previously established for lymphocytes and mononuclear phagocytes. Dendritic cells are not found in newborn mice, and their concentration in both spleen and mesenteric lymph node does not reach adult levels until 3-4 wk of age. Dendritic cells largely disappear from adherent populations following administration of steroids (2.5 mg hydrocortisone acetate s.c.) and ionizing radiation (D(o) of 100 rads for Co(60)). Splenic dendritic cells can originate from precursors located in both bone marrow and spleen itself, probably the red pulp. The mature splenic population does not actively divide (pulse labeling index with [(3)H]thymidine of 1.5-2.5%), but does turnover at substantial rate, 10+% of the total pool per day. The influx of new cells appears to be derived from a proliferating precursor compartment, but the mechanism for efflux or turnover is not known. Dendritic cells in spleen and node undergo little or moderate increase in numbers during development of a primary immune response. These in vivo characteristics, taken together, further distinguish dendritic cells as a novel cell type, distinct from mononuclear phagocytes and lymphocytes.

Keratinocytes electroporated with human papillomavirus (HPV) DNA (HPV-6, 11, 16 and 18) exhibited an increased cellular proliferation which was quantitated as microcolony and macrocolony formation. However, only macrocolonies induced by HPV-16 or HPV-18 DNA (the two viral types most commonly found in human cervical carcinomas) gave rise to proliferating, poorly-stratified colonies when grown in the presence of serum and calcium. Hydrocortisone increased the frequency of these differentiation-resistant colonies, and studies showed that they were immortalized, contained one copy of viral DNA per cell, expressed three discrete species of viral RNA and synthesized the viral E7 protein. HPV-induced cellular proliferation and altered differentiation are therefore separable events and may represent the activity of different viral genes.

Damage of the endothelial glycocalyx, which ranges from 200 to 2000 nm in thickness, decreases vascular barrier function and leads to protein extravasation and tissue oedema, loss of nutritional blood flow, and an increase in platelet and leucocyte adhesion. Thus, its protection or the restoration of an already damaged glycocalyx seems to be a promising therapeutic target both in an acute critical care setting and in the treatment of chronic vascular disease. Drugs that can specifically increase the synthesis of glycocalyx components, refurbish it, or selectively prevent its enzymatic degradation do not seem to be available. Pharmacological blockers of radical production may be useful to diminish the oxygen radical stress on the glycocalyx. Tenable options are the application of hydrocortisone (inhibiting mast-cell degranulation), use of antithrombin III (lowering susceptibility to enzymatic attack), direct inhibition of the cytokine tumour necrosis factor-alpha, and avoidance of the liberation of natriuretic peptides (as in volume loading and heart surgery). Infusion of human plasma albumin (to maintain mechanical and chemical stability of the endothelial surface layer) seems the easiest treatment to implement.

The term ''adrenergic'' originates from ''adrenaline'' and describes hormones or drugs whose effects are similar to those of epinephrine. Adrenergic stress is mediated by stimulation of adrenergic receptors and activation of post-receptor pathways. Critical illness is a potent stimulus of the sympathetic nervous system. It is undisputable that the adrenergic-driven ''fight-flight response'' is a physiologically meaningful reaction allowing humans to survive during evolution. However, in critical illness an overshooting stimulation of the sympathetic nervous system may well exceed in time and scope its beneficial effects. Comparable to the overwhelming immune response during sepsis, adrenergic stress in critical illness may get out of control and cause adverse effects. Several organ systems may be affected. The heart seems to be most susceptible to sympathetic overstimulation. Detrimental effects include impaired diastolic function, tachycardia and tachyarrhythmia, myocardial ischemia, stunning, apoptosis and necrosis. Adverse catecholamine effects have been observed in other organs such as the lungs (pulmonary edema, elevated pulmonary arterial pressures), the coagulation (hypercoagulability, thrombus formation), gastrointestinal (hypoperfusion, inhibition of peristalsis), endocrinologic (decreased prolactin, thyroid and growth hormone secretion) and immune systems (immunomodulation, stimulation of bacterial growth), and metabolism (increase in cell energy expenditure, hyperglycemia, catabolism, lipolysis, hyperlactatemia, electrolyte changes), bone marrow (anemia), and skeletal muscles (apoptosis). Potential therapeutic options to reduce excessive adrenergic stress comprise temperature and heart rate control, adequate use of sedative/analgesic drugs, and aiming for reasonable cardiovascular targets, adequate fluid therapy, use of levosimendan, hydrocortisone or supplementary arginine vasopressin.

Sepsis is physiologically viewed as a proinflammatory and procoagulant response to invading pathogens. There are three recognized stages in the inflammatory response with progressively increased risk of end-organ failure and death: sepsis, severe sepsis, and septic shock. Patients with cirrhosis are prone to develop sepsis, sepsis-induced organ failure, and death. There is evidence that in cirrhosis, sepsis is accompanied by a markedly imbalanced cytokine response ("cytokine storm"), which converts responses that are normally beneficial for fighting infections into excessive, damaging inflammation. Molecular mechanisms for this excessive proinflammatory response are poorly understood. In patients with cirrhosis and severe sepsis, high production of proinflammatory cytokines seems to play a role in the worsening of liver function and the development of organ/system failures such as shock, renal failure, acute lung injury or acute respiratory distress syndrome, coagulopathy, or hepatic encephalopathy. In addition, these patients may have sepsis-induced hyperglycemia, defective arginine-vasopressin secretion, adrenal insufficiency, or compartmental syndrome. In patients with cirrhosis and spontaneous bacterial peritonitis (SBP), early use of antibiotics and intravenous albumin administration decreases the risk for developing renal failure and improves survival. There are no randomized studies that have been specifically performed in patients with cirrhosis and severe sepsis to evaluate treatments that have been shown to improve outcome in patients without cirrhosis who have severe sepsis or septic shock. These treatments include recombinant human activated C protein and protective-ventilation strategy for respiratory failure. Other treatments should be evaluated in the cirrhotic population with severe sepsis including the early use of antibiotics in "non-SBP" infections, vasopressor therapy, hydrocortisone, renal-replacement therapy and liver support systems, and selective decontamination of the digestive tract or oropharynx.

Colony formation by normal human epidermal keratinocytes (HK) has been achieved in a medium that contains no deliberately added undefined supplements. The term "defined" is used to describe this medium, although the possibility that trace contaminants in its components could be contributing to the multiplication that it supports cannot yet be ruled out completely. The defined medium consists of a basal medium, MCDB 152, supplemented with 5 ng/ml epidermal growth factor (EGF), 10 micrograms/ml transferrin, 5 micrograms/ml insulin, 1.4 X 10(-6) M hydrocortisone, 1.0 X 10(-5) Methanolamine, 1.0 X 10(-5) M phosphoethanolamine, and 2.0 X 10(-9) M progesterone. MCDB 152 differs from MCDB 151, previously developed for multiplication of HK with small amounts of dialyzed serum (Peehl and Ham, 1980b), only by addition of the trace element mixture from human fibroblast medium MCDB 104 (McKeehan et al., 1977). Most of the requirement for transferrin, which is the least defined component of the defined medium, can be replaced by adding freshly dissolved and sterilized ferrous sulfate to the final medium after it has been filter sterilized. Insulin and EGF are clearly needed for optimal multiplication and hydrocortisone is mildly beneficial. Either ethanolamine or phosphoethanolamine must be present in the defined medium for HK multiplication. There is a greater need for EGF and less for hydrocortisone in the defined medium than in previous partially defined systems that we have worked with. Very large colonies of flattened epithelial cells are obtained in the defined medium, which has a low calcium concentration (0.03 mM) and does not favor keratinocyte differentiation. Less growth and more differentiation are obtained with higher calcium concentrations. The defined medium is highly selective for keratinocyte growth from a mixed inoculum of keratinocytes and fibroblasts.

The purpose of this study was to identify culture conditions for maintenance of isolated mouse type II cells with intact surfactant protein (SP) and phospholipid production. Type II cells were isolated from 6-wk-old mice and cultured on Matrigel matrix-rat tail collagen (70:30 vol/vol) in bronchial epithelial cell growth medium minus hydrocortisone plus 5% charcoal-stripped FBS and 10 ng/ml keratinocyte growth factor. Under these conditions, type II cells actively produced surfactant phospholipids and proteins for at least 7 days. Synthesis and secretion of surfactant phospholipids and SP-A, -B, -C, and -D declined on day 1 of culture but recovered by day 3, reaching levels comparable to or exceeding freshly isolated cells by day 5. Abundant lamellar bodies were readily apparent in cells examined on days 5 and 7, and a surfactant pellet was recovered by centrifugation of media harvested on each day of culture. Secretion of SP-B, SP-C, and phosphatidylcholine was stimulated by phorbol 12-myristate 13-acetate and was inhibited by compound 48/80. When tested with a bubble surfactometer, surfactant secreted by type II cells on day 5 of culture lowered surface tension to 5.2 +/- 2.3 mN/m. This is the first description of the synthesis and secretion of a functional surfactant complex by mouse type II cells after 7 days in primary culture.

OBJECTIVE

Autoimmune disease (e.g., Cogan syndrome) and other inflammatory inner ear diseases may ravage the labyrinth if not treated aggressively with antiinflammatory medication. Corticosteroids are the mainstay of treatment, yet, partly because of the existence of the blood-labyrinthine barrier, the ideal drug, dose, and route of administration are currently unknown.

METHODS

In the present study, we established cochlear fluid pharmacokinetic profiles of hydrocortisone, methylprednisolone, and dexamethasone in the guinea pig following oral, intravenous, and topical (intratympanic) administration. High-performance liquid chromatography was used to determine the drug concentrations, and comparisons were made with simultaneous pharmacokinetic profiles from blood and cerebrospinal fluid.

RESULTS

Our findings demonstrated a much higher penetration of all three drugs into the cochlear fluids following topical application as compared with systemic administration, with methylprednisolone showing the best profile.

CONCLUSIONS

The results suggested that intratympanic administration of corticosteroids might be more efficacious while avoiding high blood levels and therefore the deleterious side effects of systemic use.

METHODS

Thirty-seven patients with various inner ear disorders causing sensorineural hearing loss were subsequently treated using intratympanic corticosteroids, 20 with dexamethasone, and 17 with methlyprednisolone. Patients with immune-mediated hearing losses showed the best results, with notable improvement also seen in several cases of a "sudden deafness." No benefit was seen in patients with cochlear hydrops or those with sudden deterioration of a preexisting hearing loss. Three patients developed a transient otitis media related to the treatments, easily controlled with antibiotics. There were no cases of treatment-induced hearing loss and no permanent tympanic membrane perforations.

CONCLUSIONS

Overall injection of intratympanic corticosteroids for the treatment of hearing loss in inner ear disorders appears to be both safe and highly effective for certain disorders. The concept of this technique is supported by animal experimental data. The findings from the present study warrant further clinical application and experimental investigation.

BACKGROUND

A variety of interventions have been used in the treatment and management of chronic fatigue syndrome (CFS). Currently, debate exists among health care professionals and patients about appropriate strategies for management.

OBJECTIVE

To assess the effectiveness of all interventions that have been evaluated for use in the treatment or management of CFS in adults or children.

METHODS

Nineteen specialist databases were searched from inception to either January or July 2000 for published or unpublished studies in any language. The search was updated through October 2000 using PubMed. Other sources included scanning citations, Internet searching, contacting experts, and online requests for articles.

METHODS

Controlled trials (randomized or nonrandomized) that evaluated interventions in patients diagnosed as having CFS according to any criteria were included. Study inclusion was assessed independently by 2 reviewers. Of 350 studies initially identified, 44 met inclusion criteria, including 36 randomized controlled trials and 8 controlled trials.

METHODS

Data extraction was conducted by 1 reviewer and checked by a second. Validity assessment was carried out by 2 reviewers with disagreements resolved by consensus. A qualitative synthesis was carried out and studies were grouped according to type of intervention and outcomes assessed.

RESULTS

The number of participants included in each trial ranged from 12 to 326, with a total of 2801 participants included in the 44 trials combined. Across the studies, 38 different outcomes were evaluated using about 130 different scales or types of measurement. Studies were grouped into 6 different categories. In the behavioral category, graded exercise therapy and cognitive behavioral therapy showed positive results and also scored highly on the validity assessment. In the immunological category, both immunoglobulin and hydrocortisone showed some limited effects but, overall, the evidence was inconclusive. There was insufficient evidence about effectiveness in the other 4 categories (pharmacological, supplements, complementary/alternative, and other interventions).

CONCLUSIONS

Overall, the interventions demonstrated mixed results in terms of effectiveness. All conclusions about effectiveness should be considered together with the methodological inadequacies of the studies. Interventions which have shown promising results include cognitive behavioral therapy and graded exercise therapy. Further research into these and other treatments is required using standardized outcome measures.

Respiratory morbidity including respiratory distress syndrome (RDS) is a serious complication of preterm birth and the primary cause of early neonatal mortality and disability. While researching the effects of the steroid dexamethasone on premature parturition in fetal sheep in 1969, Liggins found that there was some inflation of the lungs of lambs born at gestations at which the lungs would be expected to be airless. Liggins and Howie published the first randomised controlled trial in humans in 1972 and many others followed.

To assess the effects of administering a course of corticosteroids to the mother prior to anticipated preterm birth on fetal and neonatal morbidity and mortality, maternal mortality and morbidity, and on the child in later life.

We searched Cochrane Pregnancy and Childbirth's Trials Register (17 February 2016) and reference lists of retrieved studies.

We considered all randomised controlled comparisons of antenatal corticosteroid administration (betamethasone, dexamethasone, or hydrocortisone) with placebo, or with no treatment, given to women with a singleton or multiple pregnancy, prior to anticipated preterm delivery (elective, or following spontaneous labour), regardless of other co-morbidity, for inclusion in this review. Most women in this review received a single course of steroids; however, nine of the included trials allowed for women to have weekly repeats.

Two review authors independently assessed trials for inclusion and risk of bias, extracted data and checked them for accuracy. The quality of the evidence was assessed using the GRADE approach.

This update includes 30 studies (7774 women and 8158 infants). Most studies are of low or unclear risk for most bias domains. An assessment of high risk usually meant a trial had potential for performance bias due to lack of blinding. Two trials had low risks of bias for all risk of bias domains.Treatment with antenatal corticosteroids (compared with placebo or no treatment) is associated with a reduction in the most serious adverse outcomes related to prematurity, including: perinatal death (average risk ratio (RR) 0.72, 95% confidence interval (CI) 0.58 to 0.89; participants = 6729; studies = 15; Tau² = 0.05, I² = 34%; moderate-quality); neonatal death (RR 0.69, 95% CI 0.59 to 0.81; participants = 7188; studies = 22), RDS (average RR 0.66, 95% CI 0.56 to 0.77; participants = 7764; studies = 28; Tau² = 0.06, I² = 48%; moderate-quality); moderate/severe RDS (average RR 0.59, 95% CI 0.38 to 0.91; participants = 1686; studies = 6; Tau² = 0.14, I² = 52%); intraventricular haemorrhage (IVH) (average RR 0.55, 95% CI 0.40 to 0.76; participants = 6093; studies = 16; Tau² = 0.10, I² = 33%; moderate-quality), necrotising enterocolitis (RR 0.50, 95% CI 0.32 to 0.78; participants = 4702; studies = 10); need for mechanical ventilation (RR 0.68, 95% CI 0.56 to 0.84; participants = 1368; studies = 9); and systemic infections in the first 48 hours of life (RR 0.60, 95% CI 0.41 to 0.88; participants = 1753; studies = 8).There was no obvious benefit for: chronic lung disease (average RR 0.86, 95% CI 0.42 to 1.79; participants = 818; studies = 6; Tau² = 0.38 I² = 65%); mean birthweight (g) (MD -18.47, 95% CI -40.83 to 3.90; participants = 6182; studies = 16; moderate-quality); death in childhood (RR 0.68, 95% CI 0.36 to 1.27; participants = 1010; studies = 4); neurodevelopment delay in childhood (RR 0.64, 95% CI 0.14 to 2.98; participants = 82; studies = 1); or death into adulthood (RR 1.00, 95% CI 0.56 to 1.81; participants = 988; studies = 1).Treatment with antenatal corticosteroids does not increase the risk of chorioamnionitis (RR 0.83, 95% CI 0.66 to 1.06; participants = 5546; studies = 15; moderate-quality evidence) or endometritis (RR 1.20, 95% CI 0.87 to 1.63; participants = 4030; studies = 10; Tau² = 0.11, I² = 28%; moderate-quality). No increased risk in maternal death was observed. However, the data on maternal death is based on data from a single trial with two deaths; four other trials reporting maternal death had zero events (participants = 3392; studies = 5; moderate-quality).There is no definitive evidence to suggest that antenatal corticosteroids work differently in any pre-specified subgroups (singleton versus multiple pregnancy; membrane status; presence of hypertension) or for different study protocols (type of corticosteroid; single course or weekly repeats).GRADE outcomes were downgraded to moderate-quality. Downgrading decisions (for perinatal death, RDS, IVH, and mean birthweight) were due to limitations in study design or concerns regarding precision (chorioamnionitis, endometritis). Maternal death was downgraded for imprecision due to few events.

Evidence from this update supports the continued use of a single course of antenatal corticosteroids to accelerate fetal lung maturation in women at risk of preterm birth. A single course of antenatal corticosteroids could be considered routine for preterm delivery. It is important to note that most of the evidence comes from high income countries and hospital settings; therefore, the results may not be applicable to low-resource settings with high rates of infections.There is little need for further trials of a single course of antenatal corticosteroids versus placebo in singleton pregnancies in higher income countries and hospital settings. However, data are sparse in lower income settings. There are also few data regarding risks and benefits of antenatal corticosteroids in multiple pregnancies and other high-risk obstetric groups. Further information is also required concerning the optimal dose-to-delivery interval, and the optimal corticosteroid to use.We encourage authors of previous studies to provide further information, which may answer any remaining questions about the use of antenatal corticosteroids in such pregnancies without the need for further randomised controlled trials. Individual patient data meta-analysis from published trials is likely to answer some of the evidence gaps. Follow-up studies into childhood and adulthood, particularly in the late preterm gestation and repeat courses groups, are needed. We have not examined the possible harmful effects of antenatal corticosteroids in low-resource settings in this review. It would be particularly relevant to explore this finding in adequately powered prospective trials.

Transplantation studies demonstrate that an epithelial stem cell component must exist in the mouse mammary gland throughout life. Samples taken from any portion of the mammary gland at any age and at any developmental stage, including full functional differentiation, give rise to mammary epithelial outgrowths with complete developmental capacity. Cytological examination of mouse mammary gland explants revealed the presence of morphologically distinct cells distributed sporadically among the mammary epithelium, whose behaviour in vivo and in vitro suggested that they might represent a latent epithelial stem cell population. These pale-staining cells possessed large spherical nuclei, a clear cytoplasm and a round smooth-contoured shape. Electron microscopy confirmed their pale-staining characteristics and revealed a cytoplasm sparsely populated with organellar structures, such as mitochondria and endoplasmic reticulum. Their epithelial genealogy was demonstrated by the presence of terminal bars and tight junctions formed with their epithelial cell neighbours. In vivo, these cells were found among mammary epithelial cell populations in 16-day-old embryos onward in both ductal or lobular structures during all stages of pregnancy, lactation and involution. In explant cultures, these cells did not undertake a secretory morphology in the presence of lactogenic hormones, although occasionally they became immunologically positive for casein. They did not incorporate [3H]-thymidine into their nuclei under any of the experimental conditions used; however, they appeared to undergo mitosis within 4 h regardless of the presence or absence of hormone(s). At 24 h increased numbers of pale cells were found in pairs or in groups. At 72 h in the presence of IFPrl (medium containing insulin, hydrocortisone and prolactin), the pairs and groups of pale cells observed at 24 h were not found. Instead, individual pale cells were seen among groups of cytologically and functionally differentiated secretory epithelial cells. When lactogenic hormones were not present, groups of pale cells were still present in the explants at 72 h. These findings suggest that the pale cells are arrested at G2 phase of the cell cycle and that they give rise by mitosis to daughter cells capable of differentiating in the presence of lactogenic stimuli. Inhibition of DNA synthesis in the explants did not alter these cellular events.

BACKGROUND

No consensus exists for management of adults with congenital adrenal hyperplasia (CAH) due to a paucity of data from cohorts of meaningful size.

OBJECTIVE

Our objective was to establish the health status of adults with CAH.

METHODS

We conducted a prospective cross-sectional study of adults with CAH attending specialized endocrine centers across the United Kingdom.

METHODS

Participants included 203 CAH patients (199 with 21-hydroxylase deficiency): 138 women, 65 men, median age 34 (range 18-69) years.

METHODS

Anthropometric, metabolic, and subjective health status was evaluated. Anthropometric measurements were compared with Health Survey for England data, and psychometric data were compared with appropriate reference cohorts.

RESULTS

Glucocorticoid treatment consisted of hydrocortisone (26%), prednisolone (43%), dexamethasone (19%), or a combination (10%), with reverse circadian administration in 41% of patients. Control of androgens was highly variable with a normal serum androstenedione found in only 36% of patients, whereas 38% had suppressed levels suggesting glucocorticoid overtreatment. In comparison with Health Survey for England participants, CAH patients were significantly shorter and had a higher body mass index, and women with classic CAH had increased diastolic blood pressure. Metabolic abnormalities were common, including obesity (41%), hypercholesterolemia (46%), insulin resistance (29%), osteopenia (40%), and osteoporosis (7%). Subjective health status was significantly impaired and fertility compromised.

CONCLUSIONS

Currently, a minority of adult United Kingdom CAH patients appear to be under endocrine specialist care. In the patients studied, glucocorticoid replacement was generally nonphysiological, and androgen levels were poorly controlled. This was associated with an adverse metabolic profile and impaired fertility and quality of life. Improvements in the clinical management of adults with CAH are required.

Arachidonate 15-lipoxygenase (arachidonate:oxygen 15-oxidoreductase, EC 1.13.11.33) is a lipid-peroxidating enzyme that is implicated in oxidizing low density lipoprotein to its atherogenic form. Monocyte/macrophage 15-lipoxygenase is present in human atherosclerotic lesions. To pursue a basis for induction of the enzyme, which is not present in blood monocytes, the ability of relevant cytokines to regulate its expression was investigated. Interleukin 4 (IL-4), among 16 factors tested, specifically induced 15-lipoxygenase mRNA and protein in cultured human monocytes. Interferon gamma and hydrocortisone inhibited this induction. High-performance liquid chromatography analysis of lipid extracts from IL-4-treated monocytes detected 15-lipoxygenase products esterified to the cellular membrane lipids, indicating enzymatic action on endogenous substrates. Stimulation of IL-4-treated monocytes with calcium ionophore or opsonized zymosan A enhanced the formation of 15-lipoxygenase products. These data identify IL-4 and interferon gamma as physiological regulators of lipoxygenase expression and suggest an important link between 15-lipoxygenase function and the immune/inflammatory response in atherosclerosis as well as other diseases.

Reduction mammoplasty tissue was used to obtain short-term cultures of human epithelial cell populations. Digestion of tissue with collagenase and hyaluronidase resulted in cell clusters (organoids) resembling ductal and alveolar structures; these could be separated by filtration from the stromal components. Epithelial outgrowth from these organoids was greatly enhanced by the addition of conditioned medium from other human epithelial and myoepithelial cell lines. Additionally, the mammary epithelial growth was stimulated by insulin, hydrocortisone, epidermal growth factor, and steroid hormones. With this enriched nutritional environment, active cell division could be maintained for 1 to 3 months and cells could be serially subcultured 1 to 4 times.

Results are reported showing that pituitary extracts stimulate growth of 3T3 cells, an established line of mouse fibroblast. Cells were cultured in medium containing an organic buffer (HEPES; N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid), which increases the serum requirement for cell growth. The pituitary growth-prompting activity is (a) not dialyzable; (b) thermolabile; (c) protease sensitive; and (d) enhanced by hydrocortisone (a steroid hormone). Activity was quantitatively assayed by measurement of stimulation of cell division or [(3)H]thymidine uptake into DNA. The active species is probably different from pituitary protein hormones and it seems to be effective at concentrations observed for protein hormones in vivo. 3T3 cells transformed by simian virus 40 are not responsive to this activity.

Homeostasis of the central nervous system (CNS) microenvironment is maintained by the blood-brain barrier (BBB) which regulates the transport of molecules from blood into brain and back. Many disorders change the functionality and integrity of the BBB. Glucocorticoids are being used sucessfully in the treatment of some disorders while their effects on others are questionable. In addition, conflicting results between clinical and experimental experience using animal models has arisen, so that the results of molecular studies in animal models need to be revisited in an appropriate in vitro model of the human BBB for more effective treatment strategies. Using the human brain microvascular endothelial cell line hCMEC/D3, the influence of glucocorticoids on the expression of barrier constituting adherens junction and tight junction transmembrane proteins (VE-cadherin, occludin, claudins) was investigated and compared to other established BBB models. In hCMEC/D3 cells the administration of glucocorticoids induced expression of the targets occludin 2.75 +/- 0.04-fold and claudin-5 up to 2.32 +/- 0.11-fold, which is likely to contribute to the more than threefold enhancement of transendothelial electrical resistance reflecting barrier tightness. Our analyses further provide direct evidence that the GC hydrocortisone prevents endothelial barrier breakdown in response to pro-inflammatory stimuli (TNFalpha administration), which could be demonstrated to be partly based on maintenance of occludin levels. Our studies strongly suggest stabilization of BBB function as a mode of GC action on a molecular level in the human brain vasculature.

The vascular endothelial growth factor (VEGF) is a specific mitogen for vascular endothelial cells and enhances vascular permeability and edemagenesis. VEGF is also a major regulator of angiogenesis and may be a key target for inhibiting angiogenesis in angiogenesis-associated diseases. Among the extensively studied angiostatic compounds are several corticosteroids when used alone or in combination with heparin. In this study we present evidence for an additional mechanism of action of hydrocortisone, cortisone and dexamethasone in inhibiting edemagenesis or angiogenesis. In cultures of aortic human vascular smooth muscle cells these corticosteroids (1 x 10(-8) to 1 x 10(-12) M) abolished the platelet-derived growth factor-induced (PDGF) expression of the VEGF gene in a dose-dependent manner. In contrast, two precursors of corticosteroids, desoxycorticosterone or pregnenolone, did not affect PDGF-induced VEGF expression. Our findings indicate that the capacity of corticosteroids to reduce edema or to prevent new blood vessel formation may be attributed, at least in part to the ability of these agents to abolish the expression of VEGF.

Human mesothelial cells grew rapidly in culture when provided with serum, EGF, and hydrocortisone, adopting a fibroblastoid shape and forming parallel, multilayered arrays at saturation density. In the absence of EGF, the cells grew slowly to a flat, epithelioid monolayer similar to their normal pattern in vivo. Mesothelial cells normally have a high keratin and a low vimentin content in vivo. In culture, rapidly growing cells greatly reduced synthesis and content of their four major keratins to levels undetectable by immunofluorescence in most cells, but keratin synthesis and content returned to high levels whenever growth slowed. Vimentin synthesis and content was high during serial culture, but decreased several-fold in nondividing cells. The unique ability of the mesothelial cell to reversibly alter its morphology and intermediate filament composition is of unknown function and mechanism, but accounts for the morphological heterogeneity and the presence of keratin-negative cells in mesotheliomas.

Forty-four patients with documented meningeal carcinomatosis (small-cell lung carcinoma [SCLC], 29%; breast carcinoma, 25%) were treated in a prospective randomized trial with intrathecal methotrexate (MTX) 15 mg or MTX plus cytosine arabinoside (Ara-C) 50 mg/m2. Most patients received intrathecal hydrocortisone (HC) each treatment to minimize arachnoiditis. Overall response was 55%. Seven patients achieved complete response. Response to MTX was superior to combined MTX/Ara-C, but not significantly so (61% v 45%; P greater than .10). Response was more frequent if drugs were administered via Ommaya reservoir than by lumbar puncture (65% v 48%; P greater than .10). Concurrent radiotherapy to the CNS was associated with significantly better response (73% v 35%; P less than .05). Small-cell lung carcinoma patients showed the best response (69%). Overall median survival for the whole group was 8 weeks, but responders fared better than nonresponders (median survival, 18 v 7 weeks; P less than .05). Nausea and vomiting were the most common toxicities encountered (45%), but rarely proved limiting. An unusual, previously undocumented reaction to intrathecal HC was noted. MTX is moderately effective in nonleukemic meningeal carcinomatosis, but the addition of Ara-C does not appear to improve results. Pretreatment factors did not predict outcome in this trial.

IL-6 is a regulator of inflammatory and immunological processes and high levels of this cytokine have recently been shown to be present in synovial fluids from inflammatory and degenerative arthropathies. Synovial fluid IL-6 may in part be a product of synoviocytes that have previously been identified as a source of IL-6. To further define intraarticular sources of IL-6, we examined the ability of chondrocytes to produce IL-6 and studied its modulation by inflammatory cytokines and homeostatic regulators of chondrocyte function. Human articular chondrocytes isolated from normal and osteoarthritic joints released low levels of IL-6 when cultured in the presence of serum. The inflammatory cytokines IL-1, and, to a lesser extent, TNF-alpha and IFN-gamma increased IL-6 production. Among the growth factors known to act on chondrocytes, only transforming growth factor-beta, but not epidermal growth factor. fibroblast growth factor, insulin growth factor-1 and 2, platelet growth factor or insulin, was able to significantly increase IL-6 synthesis. Analysis of hormonal influences on chondrocyte IL-6 production showed that testosterone and estradiol synergized with IL-1 in the induction of IL-6. Hydrocortisone, at 10 ng/ml, reduced IL-1-induced IL-6 production by more than 50%. Chondrocyte-derived IL-6 stimulated acute phase protein synthesis and hybridoma cell proliferation. These biological activities were neutralized by a specific antibody to IL-6. Metabolic labeling and immunoprecipitation studies showed that IL-1 induced de novo synthesis of IL-6 and that the IL-6 proteins secreted by chondrocytes were similar to those from fibroblasts. These results demonstrate that chondrocytes are able to produce IL-6 in response to physiologic and inflammatory stimuli. Chondrocytes probably contribute to the increased synovial fluid levels of IL-6 in inflammatory and degenerative conditions of cartilage, and IL-6 may serve as a mediator to coordinate responses to cartilage injury.

Quantitative stereological methods have been adapted for the measurement of the volume of liver attributable to parenchymal, hematopoietic, and Kupffer cells and for the measurement of the relative and absolute number (per unit volume) of these cell types and the mean volume of the parenchymal cell. These morphological parameters are the main ones for interpreting the biochemical differentiation of liver. Quantitative changes in these parameters, in rat liver between the 15th day of gestation and adult life, are presented. Despite the large number of hematopoietic cells, the parenchymal cells fill more than half of the liver volume between the 15th and 18th days of gestation and 0.85 of the liver volume at term. The fraction of liver volume occupied by Kupffer cells is never more than 0.02; the number of Kupffer cells per cubic centimeter increases less than twofold between fetal and adult life. The mean volume of individual parenchymal cells undergoes a threefold rise during late fetal life, declines in the neonatal period, and doubles between the 12th and 28th postnatal days. With the morphometric data obtained, it is impossible to convert enzyme concentrations (units per gram, determined in homogenates of whole liver) to enzyme amounts per unit volume of parenchymal or hematopoietic tissue or per individual cell of either type. In late fetal liver, only rises in enzyme concentration less than twofold may be attributed to the enrichment of parenchymal tissue at the expense of hematopoietic elements. The sudden upsurge, by more than twofold, of hepatic enzymes of the late fetal cluster (and also of the neonatal and late suckling cluster) reflects rises per parenchymal mass and per parenchymal cell. Thyroxine and glucagon, the administration of which to fetal rats promotes enzyme differentiation in liver, are without appreciable effect on the cytological parameters studied. Hydrocortisone accelerates the involution of hematopoietic tissue in fetal liver. Enzymes that are diminished by prenatal injection of hydrocortisone may be concentrated in hematopoietic cells.