The effects of insulin on basal and hydrocortisone-induced growth hormone (GH) secretion were studied in rat pituitary tumor cells (GH3). Cells were grown in monolayer culture and exposed to exogenously added insulin for up to 8 d. Basal GH secretion was inhibited by insulin (0.7 nM) after a 48-h lag period by approximately 50% (P less than 0.01, vs. untreated control cells). The suppression of GH secretion was reversible, as removal of added insulin resulted in return of GH secretion to normal levels after 24 h. Maximal suppression of basal GH secretion was achieved by 0.7 nM insulin, and these effects were prevented by simultaneous exposure of the cells to guinea pig anti-insulin serum (1:2,000). No effects of insulin on cell replication were evident, and glucose concentration in the medium did not differ in control or insulin-treated wells. Insulin (7 nM) significantly suppressed the fivefold hydrocortisone-induced GH stimulation during 5 d of incubation with up to 1,000 nM of the steroid (P less than 0.001). These inhibitory effects were similarly observed in glucose- and pyruvate-free medium, and in the presence of 2-deoxyglucose. Insulin also reversed the suppression of prolactin (PRL) secretion induced by hydrocortisone (1 uM), and actually stimulated basal PRL secretion by over 50%. Insulin did not alter the inhibitory effect of hydrocortisone on GH3 cell proliferation. Although higher doses (13 nM) of insulin-like growth factor (IGF-I) also suppressed basal GH secretion, IGF-I did not alter the GH and PRL secretory changes induced by hydrocortisone. The results show that insulin exerts a direct, specific inhibitory effect on basal and hydrocortisone-induced GH secretion by GH3 cells unrelated to glucose utilization by the cells.

The kinetics of manganese(II) ion uptake and efflux have been investigated using tracer 54Mn(II) with glial cells cultured from chick cerebral cortex in chemically defined medium. The initial velocity of Mn(II) uptake versus [Mn(II)] exhibit saturation, with an apparent S0.5 approximately 18(+/- 3) microM. Both the rate and extent of Mn(II) uptake are inhibited by Ca(II), either added externally or preloaded into the glial cells. Preloading of glia with Mn(II) also inhibits the rate of external 54Mn(II) uptake. Zn(II) inhibits but Cu(II) activates Mn(II) uptake. Efflux of Mn(II) from preloaded cells occurs as a biphasic process, with rapid release of 30-40% of total cell Mn(II), then much slower release of the remainder. Permeabilization of cells with dextran sulfate also rapidly released ca. 30% of total cell Mn(II). High external Mn(II) enhanced both the rate and extent of Mn(II) efflux. CCCP, an uncoupler of oxidative phosphorylation, inhibited both Mn(II) uptake and efflux significantly, but addition of cyanide, ouabain, insulin, hydrocortisone, K+, or Nd(III) had no effect on either process. Taken together, these data suggest a model in which Mn(II) is brought across the plasma membrane by facilitated diffusion, binds to cytosolic protein sites, and is partitioned into the mitochondria by an active transport mechanism. The fact that the Mn(II) flux rates observed with cultured glia are much faster than those reported for overall uptake and efflux of brain Mn(II) in vivo suggests that the blood-brain barrier may play a significant role in determining these latter rates in whole animals.

Cortisol measurements in blood, saliva and urine are frequently used to examine the hypothalamus-pituitary-adrenal (HPA) axis in clinical practice and in research. However, cortisol levels are subject to variations due to acute stress, the diurnal rhythm and pulsatile secretion. Cortisol measurements in body fluids are not always a reflection of long-term cortisol exposure. The analysis of cortisol in scalp hair is a relatively novel method to measure cumulative cortisol exposure over months up to years. Over the past years, hair cortisol concentrations (HCC) have been examined in association with a large number of somatic and mental health conditions. HCC can be used to evaluate disturbances of the HPA axis, including Cushing's syndrome, and to evaluate hydrocortisone treatment. Using HCC, retrospective timelines of cortisol exposure can be created which can be of value in diagnosing cyclic hypercortisolism. HCC have also been shown to increase with psychological stressors, including major life events, as well as physical stressors, such as endurance exercise and shift work. Initial studies show that HCC may be increased in depression, but decreased in general anxiety disorder. In posttraumatic stress disorder, changes in HCC seem to be dependent on the type of traumatic experience and the time since traumatization. Increased hair cortisol is consistently linked to obesity, metabolic syndrome and cardiovascular disease. Potentially, HCC could form a future marker for cardiovascular risk stratification, as well as serve as a treatment target.

BACKGROUND

Conventional hydrocortisone dosing schedules do not mimic the normal circadian rhythm of cortisol, making it difficult to optimize treatment in congenital adrenal hyperplasia (CAH).

METHODS

We report a 14.5-year-old boy with CAH who had reduced bioavailability [42% (normal 80% orally and 100% by im route)] and increased clearance [half-life 50 min (normal range, 70-100 min)] of oral doses of hydrocortisone leading to ambient serum 17-hydroxyprogesterone concentrations of 400 nmol/liter (14.5 ng/ml) and androstenedione concentrations of 24.9 nmol/liter (7.1 ng/ml).

METHODS

Using a continuous but variable sc hydrocortisone infusion via an insulin pump, rapid control of his CAH was attained with a normal cortisol circadian profile. Average daily hydrocortisone dose was 17.4-18.6 mg/m(2), which produces on average 24-h serum cortisol and 17-hydroxyprogesterone concentrations of 316 nmol/liter (115 ng/ml) and 4.3 nmol/liter (1.4 ng/ml), respectively. Therapy has been maintained over 4 yr with suppression of normal adrenal androgen production and normal progression through puberty.

CONCLUSIONS

Continuous sc infusion of hydrocortisone may prove a valuable adjunct to therapy for CAH, particularly in patients requiring high doses of oral hydrocortisone and in those with abnormal hydrocortisone pharmacokinetics.

BACKGROUND

Treatment of congenital adrenal hyperplasia (CAH) is suboptimal. Inadequate suppression of androgens and glucocorticoid excess are common and current glucocorticoid formulations cannot replace the cortisol circadian rhythm.

OBJECTIVE

The primary objective was to characterize the pharmacokinetic profile of Chronocort, a modified-release hydrocortisone formulation, in adults with CAH. Secondary objectives included examining disease control following 6 months of Chronocort with dose titration.

METHODS

Sixteen adults (eight females) with classic CAH participated in an open-label, nonrandomized, Phase 2 study at the National Institutes of Health Clinical Center. Twenty-four-hour blood sampling was performed on conventional glucocorticoids and following 6 months of Chronocort. Chronocort was initiated at 10 mg (0700 h) and 20 mg (2300 h). Dose titration was performed based on androstenedione and 17-hydroxyprogresterone (17-OHP) levels and clinical symptomatology.

METHODS

The primary outcome was cortisol pharmacokinetics of Chronocort and secondary outcomes included biomarkers of CAH control (androstenedione and 17-OHP).

RESULTS

In patients with CAH, Chronocort cortisol profiles were similar to physiologic cortisol secretion. Compared with conventional therapy, 6 months of Chronocort resulted in a decrease in hydrocortisone dose equivalent (28 ± 11.8 vs 25.9 ± 7.1 mg/d), with lower 24-hour (P = .004), morning (0700-1500 h; P = .002), and afternoon (1500-2300 h; P = .011) androstenedione area under the curve (AUC) and lower 24-hour (P = .023) and morning (0700-1500 h; P = .02) 17-OHP AUC.

CONCLUSIONS

Twice-daily Chronocort approximates physiologic cortisol secretion, and was well tolerated and effective in controlling androgen excess in adults with CAH. This novel hydrocortisone formulation represents a new treatment approach for patients with CAH.

The association between inflammation of the eyes and the intestine is not often recognized by ophthalmologists. We report two patients who developed peripheral corneal ulcers, episcleritis, and scleritis just prior to the onset of Crohn's disease. The severity of the eye disease paralleled that of the intestinal symptoms, and both conditions subsided after treatment with topical steroids, oral prednisone, oral sulfasalazine, and hydrocortisone retention enemas. Inflammatory bowel disease should always be included in the differential diagnosis of scleritis and uveitis, as the patient may be benefited greatly by appropriate, early therapy of this gastrointestinal disorder.

OBJECTIVE

To investigate the effect of hydrocortisone treatment on survival without bronchopulmonary dysplasia (BPD) and to study whether serum cortisol concentrations predict the response.

METHODS

We performed a randomized, placebo-controlled trial on infants with gestation < or =30 weeks, body weight of 501 to 1250 g, and respiratory failure. Hydrocortisone was started before 36 hours of age and given for 10 days at doses from 2.0 to 0.75 mg/kg per day. Shortly before hydrocortisone treatment, basal and stimulated (ACTH, 0.1 microg/kg) serum cortisols were measured.

RESULTS

The study was discontinued early, because of gastrointestinal perforations in the hydrocortisone group (4/25 vs 0/26, P = .05); 3 of the 4 had received indomethacin/ibuprofen. The incidence of BPD (28% vs placebo 42%, P = 0.28) tended to be lower, and patent ductus arteriosus (36% vs 73%, P = .01) was lower in the hydrocortisone group. The hydrocortisone-treated infants with serum cortisol concentrations above the median had a high risk of gastrointestinal perforation. In infants with cortisol values below the median, hydrocortisone treatment increased survival without BPD.

CONCLUSIONS

Serum cortisol concentrations measured shortly after birth may identify those very high-risk infants who may benefit from hydrocortisone supplementation.

OBJECTIVE

To evaluate bone mineral density (BMD) in prepubertal and in adolescent and young adult patients with the salt-wasting form of congenital adrenal hyperplasia (CAH).

METHODS

A relationship between bone mineral content and risk for osteoporotic fractures has been observed in adulthood. Infancy, childhood, and adolescence are critical periods for skeletal mineralization; thus, chronic diseases may impair bone mass peaking, particularly if children and adolescents are overexposed to glucocorticoids, as may occur in patients with CAH. Lumbar L2-L4 BMD values were measured by dual x-ray absorptiometry and compared with those of 471 age- and sex-matched controls.

METHODS

Thirty-three patients with the salt-wasting form of CAH were studied. Sixteen (10 girls and 6 boys; age range, 1.5 to 8.3 years) were prepubertal and 17 (13 women and 4 men; age range, 17.1 to 28.2 years) were adolescent and young adults who had reached final height and had presented normal pubertal development and normal gonadal function thereafter. The average doses of hydrocortisone (mg/m body surface/day) received from diagnosis in the neonatal period to BMD evaluation were 21.2 +/- 2.2 and 22.3 +/- 2.6, respectively.

RESULTS

Mean BMD Z score values were 0.16 +/- 1.01 in prepubertal patients and 0.06 +/- 1.02 in adolescent and young adult patients with no statistically significant differences with age- and sex-matched controls. Mean height Z score values were -0.03 +/- 1.13 in prepubertal patients and -1.13 +/- 0.62 in adolescent and young adult patients with significant differences between the latter and their respective age- and sex-matched controls.

CONCLUSIONS

Long-term glucocorticoid therapy does not impair bone mass peaking in CAH patients with normal gonadal function, even though their adult height values are low.

Several aspects of in-vitro cell growth and protein synthesis were assessed in cultures of skin fibroblasts from subjects with juvenile-onset diabetes mellitus (JODM) or adult-onset diabetes mellitus (AODM) and from age-matched nondiabetic controls (C). There was an inverse correlation between increasing age and both the log-phase doubling rate and saturation density at confluence in C fibroblasts. JODM and AODM cells had a reduction in both indices of cell population growth in comparison with age-matched C fibroblasts. Fibroblasts grown in the presence of 0.3 micronM hydrocortisone were stimulated to grow more rapidly and to a greater saturation density. Stimulation of cell division by hydrocortisone accentuated the abnormalities in growth of JODM and AODM fibroblasts. Total protein and collagen synthesis was measured whtn the fibroblasts had grown to confluency in medium with or without hydrocorticone. Hydrocorticone did not produce a significant change in total protein and collagen synthesis per cell by C fibroblasts. Fibroblasts from AODM had a 180 per cent increase in total protein and collagen synthesis in the presence of hydrocortisone. In contrast, total protein and collagen synthesis decreased 40 per cent in fibroblasts from JODM when grown in the hydrocortisone medium. These studies indicate that skin fibroblast cultures from patients with diabetes exhibit abnormalities in cell proliferation. Furthermore, hydrocortisone appears to unmask diffeerences in protein synthesis that distinguish JODM and AODM fibroblasts in culture.

Mus dunni endogenous virus (MDEV) can be activated from M. dunni cells by exposing the cells to hydrocortisone or 5-iodo-2'-deoxyuridine. Interference analysis has revealed that MDEV uses a receptor for cell entry that is different from those used by other murine retroviruses. The entire genome has now been sequenced, revealing a long terminal repeat (LTR)-gag-pol-env-LTR structure typical of simple retroviruses of the murine leukemia virus genus, with no additional open reading frames between env and the 3' LTR. The LTRs and other noncoding regions of MDEV are most closely related to those of VL30 elements, while the majority of the coding sequences are most closely related to those of gibbon ape leukemia virus. MDEV represents the first example of a naturally occurring, replication-competent virus with sequences closely related to VL30 elements. The U3 region of MDEV contains six nearly perfect 80-bp repeats and the beginning of a seventh, and the region expected to contain the packaging sequence contains approximately four imperfect 33-bp repeats. The receptor specificity domains of the envelope are unique among retroviruses and show no apparent similarity to regions of known proteins.

Organic cation transporters (OCT1-3) play an important role in renal elimination of many drugs. The goals of this study were to 1) identify a cell culture model which constitutively expressed OCT2 that could be used to study the characteristics and regulation of this transporter, and 2) to study the mechanisms by which xenobiotics and hormones regulate the activity of OCT2. We characterized the endogenous organic cation transporter (OCT) activity in Madin-Darby canine kidney (MDCK) cells. The activity was localized to the basolateral membrane and was pH and membrane potential-dependent. The uptake of the model organic cation, tetraethylammonium, was saturable (Km, 19.5 +/- 4.6 microM; Vmax, 350 +/- 19.4 pmol/mg of protein/10 min) and was inhibited by known OCT inhibitors (e.g., cimetidine and quinidine). A cDNA fragment (711 base pairs) isolated by reverse transcriptase-polymerase chain reaction (RT-PCR) was greater than 83% identical to OCT2 cDNAs from mammalian species; no OCT1 or OCT3 was detected by RT-PCR, suggesting that OCT2 may be the primary basolateral OCT in MDCK. OCT2 mRNA levels were increased significantly following exposure of MDCK to the steroid hormones, dexamethasone (2.0-fold), hydrocortisone (2.4-fold), and testosterone (1.8-fold) with comparable increases in activity. Other compounds tested, including the cytochrome P450 inducers, rifampicin, phenobarbital, and phenytoin, and the OCT substrates, verapamil and metformin, had no inducing effects. Collectively, these data indicate that MDCK can serve as a useful and convenient tool in screening candidate drugs for interaction with OCT2 and for studying the regulation of this transporter. Furthermore, our data demonstrate that steroid hormones induce the transcription of OCT2 in the kidney.

Epithelial cells derived from bovine pancreatic duct have been grown continuously in culture for 30 weeks (approximately 90 doublings of the cell population). The cells were grown in Eagle's minimal essential medium supplemented with 10% heat-inactivated fetal bovine serum, 2 mM glutamine, 0.1 mM nonessential amino acids, and antibiotics. In confluent cultures, the cells are multilayered and form circular structures. When tested at various passages, the cells neither formed colonies in soft agar nor produced tumors after inoculation into athymic, nude mice. Hydrocortisone (1 and 5 microgram per ml) and insulin (1,5 and 10 microgram per ml) had no effect on the growth of the cells. beta-Retinyl acetate inhibited growth rate and cell yield at a concentration of 5 microgram per ml but was not growth-inhibitory at lower concentrations. By electron microscopy the cells have numerous mitochondria, Golgi and microvilli. Mucous droplets were observed in a small proportion of the cells. Desmosome-like structures and occluding junctions were observed more frequently between cells that had been transferred as aggregates than between cells transferred as single cells. Cytochemical studies indicated that some cells produce PAS positive granules that were not removed after treatment of the cultures with diastase. Eleven cell clones were isolated from the mass culture. The growth rates of the clones are different as well as the period of time in which the clones can be propagated in vitro.

BACKGROUND

Human mesenchymal stem cells (MSC) possess powerful ex vivo expansion and versatile differentiation potential, placing themselves at the forefront of the field of stem cell-based therapy and transplantation. Of high clinical relevance is the endothelial differentiation potential of MSC, which can be used to treat various forms of ischemic vascular disease.

METHODS

We investigated whether human umbilical cord blood (UCB)-derived MSC are able to differentiate in vitro along an endothelial lineage, by using flow cytometry, RT-PCR and immunofluorescence analyzes, as well as an Ab array method.

RESULTS

When the cells were incubated for up to 3 weeks in the presence of VEGF, EGF and hydrocortisone, they began to express a variety of endothelial lineage surface markers, such as Flk-1, Flt-1, VE-Cadherin, vWF, VCAM-1, Tie-1 and Tie-2, and to secrete a specific set of cytokines. Differentiated cells were also found to be able to uptake low-density lipoprotein and form a tubular network structure.

CONCLUSIONS

These observations have led us to conclude that UCB-derived MSC retain endothelial potential that is suitable for basic and clinical studies aimed at the development of vasculature-directed regenerative medicine.

Since the introduction of intra-articular steroid therapy 40 yr ago there have been many changes in the treatment of rheumatoid patients. Previous studies suggest differing times of response for the same agents. This study reports the response, measured by a five-point pain chart, of 300 patients with painful rheumatoid knees. Sixty received hydrocortisone succinate (HC), 150 received triamcinolone acetonide (TA), and 120 triamcinolone hexacetonide (TH). Results demonstrated little effect with HC, but good responses with TA and TH. More patients were rendered painfree for a longer time with TH; 18% at 12 weeks, as against 9% with TA (chi 2 test P < 0.005). At 12 weeks 59% showed continued improvement with TH as against 44% with TA (chi 2 test P < 0.05). TH is the preferred preparation for injection of the rheumatoid knee.

The effect of alpha- and beta-thymosin peptides, namely prothymosin alpha (ProT(alpha)), thymosin alpha(1) (T(alpha)1), parathymosin alpha (ParaT(alpha)), thymosin beta(4) (Tbeta4), thymosin beta(10) (Tbeta10), and thymosin beta(9) (Tbeta9), on the angiogenesis process was investigated using the chick chorioallantoic membrane as an in vivo angiogenesis model. The thymosin peptides tested were applied in 10 microl aliquots containing 0.01-4 nmoles of Tbeta4, Tbeta10 or Tbeta9, 0.016-6.66 nmoles of T(alpha)1, 4.1 pmoles-1.66 nmoles of ProT(alpha), and 4.4 pmoles-1.76 nmoles of ParaT(alpha). Phorbol 12-myristate 13-acetate and hydrocortisone were also used as positive and negative control, respectively. Tbeta4, ProT(alpha) and T(alpha)1 were found to enhance angiogenesis, while Tbeta10, Tbeta9 and ParaT(alpha) exhibited an inhibitory effect on the angiogenesis process. When mixtures of Tbeta4 and Tbeta10 containing active amounts of the two peptides at different proportions were applied, the promoting effect of Tbeta4 on angiogenesis was reversed in the presence of increasing concentrations of Tbeta10 and vice versa. The effect of Tbeta10, Tbeta9, ProT(alpha) and ParaT(alpha), in parallel with Tbeta4 and T(alpha)1, on the angiogenesis process was investigated for the first time as far as we know and the results of this study offer more insight into the biological regulatory roles of thymosin peptides, and provide helpful information about their therapeutic potential. Whether these agents could be used either as inhibitors of angiogenesis in disease states where uncontrolled angiogenesis is involved, e.g. in carcinogenesis, or as angiogenesis promoters that could be useful in wound healing, fracture repair, peptic ulcers etc., remains to be further studied.

The origin and kinetics of mononuclear phagocytes can only be studied after the cells have been properly characterized. The basis for such a characterization on morphological, cytochemical, functional, and immunological grounds has been discussed. The production and kinetics of mononuclear phagocytes during the normal steady state have been described and compared with the effect of an acute inflammatory stimulus on these parameters. The influence of hydrocortisone and azathioprine was also studied in this connection. The increased production and monocytosis seen during an acute inflammatory response have been shown to be regulated by a humoral factor. The concept of the mononuclear phagocyte system was also discussed and an outline of the participation of mononuclear phagocytes in pathological processes put forward.

Cells that can be cultured from pools of early-lactation milk were studied. Under the culture conditions used, the majority of cells attached to collagen-coated dishes; most of these remained single, did not divide, and in their adhesiveness, phagocytic ability, and ultrastructure resembled macrophages or histiocytes. On a plate seeded with approximately 3 X 10(5) cells, however, 10-100 colonies of dividing cells developed. These cells had the junctional complexes typical of epithelial cells and grew well in a medium supplemented with human serum and hydrocortisone for 16-20 days after seeding. After removal of serum from the medium, some cells continued to traverse the cell cycle, and the colonies containing these cells were morphologically distinct from those which became quiescent. The nondividing cells in milk could be separated from the milk epithelial cells and were able to stimulate the growth of epithelial cultures from benign mammary dysplasias.

The purpose of this study was to perform a randomized, prospective comparison of corticosteroid enemas (CS--100 mg of hydrocortisone/60 cc P.R. q.h.s.; n = 12), mesalamine enemas (5-ASA--4 g/60 cc P.R. q.h.s.; n = 19), and short-chain fatty acid enemas (SCFA--60 cc P.R. b.i.d.; n = 14) for the treatment of proctosigmoiditis. Patients presenting to the Ferguson Clinic with the diagnosis of idiopathic proctosigmoiditis were evaluated for age, sex, prior history of proctitis, duration of symptoms prior to presentation, endoscopic scoring, and mucosal biopsies. Clinical evaluation was performed at two-week intervals for six weeks, with repeat biopsies taken at six weeks. There was no significant difference with respect to age, male/female ratio, past history of proctosigmoiditis, length of colorectum involved at the time of initial presentation, symptom resolution, and endoscopic and histologic improvement among the three treatment groups. Recovery occurred in a similar proportion in each of the three groups: CS, 10/12; 5-ASA, 17/19; and SCFA, 12/14. The cost of six weeks of treatment was: CS, $71.82; 5-ASA, $347.28; and SCFA, $31.50. This study indicates that SCFA enemas are equally efficacious to CS or 5-ASA enemas for the treatment of proctosigmoiditis at a significant cost savings.

Thirty patients with distal colitis (proctosigmoiditis) in relapse were randonly allocated to twice daily treatment with traditional aqueous hydrocotrisone enemas (Cortenemas) or a suspension of hydrocortisone in an inert foam base (Colifoam). Each treatment contained the same amount of hydrocortisone. Clinical, sigmoidoscopic, and histological response was assessed after two weeks. Both agents were effective, and broadly similar in terms of objective improvement, but subjective improvement was greater with the foam preparation, and several patients expressed a preference to this mode of treatment.

1. The administration of glucagon, cAMP [adenosine 3',5'-(cyclic)-monophosphate], BcAMP [6-N-2'-O-dibutyryladenosine 3',5'-(cyclic)-monophosphate] or adrenaline to foetal rats during the last 2 days of gestation evoked the appearance of tyrosine aminotransferase and enhanced the accumulation of glucose 6-phosphatase in the liver. In foetuses 1-2 days younger only BcAMP was effective. After birth liver glucose 6-phosphatase no longer responds to glucagon or BcAMP. Tyrosine aminotransferase is still inducible by these agents in 2-day-old rats, but not in 50-day-old rats. After adrenalectomy of adults glucagon or BcAMP can enhance the induction of the enzyme by hydrocortisone. The results indicate that the ability to synthesize tyrosine aminotransferase and glucose 6-phosphatase when exposed to cAMP develops sooner than the ability to respond to glucagon with an increase in the concentration of cAMP; the responsiveness of enzymes to different hormones changes with age. A scheme illustrating the sequential development of competence in regulating the level of an enzyme is presented. 2. Actinomycin inhibited the effects of glucagon and BcAMP on liver tyrosine aminotransferase and glucose 6-phosphatase in foetal rats. Growth hormone, insulin and hydrocortisone did not enhance the formation of these enzymes. 3. The time-course of accumulation of glucose 6-phosphatase in the kidney is different from that in the liver. Hormones that increase the accumulation in foetal liver do not do so in the kidney of the same foetus or in the livers of postnatal rats.

Involucrin accumulation and ionophore-assisted envelope formation, markers of keratinocyte differentiation, were found to be highly dependent on culture conditions in the malignant epidermal keratinocyte line, SCC-13, derived from a human squamous cell carcinoma. In confluent cultures, approximately one-half of the cells were competent to form envelopes when grown in medium without hydrocortisone or retinyl acetate supplementation. Addition of hydrocortisone to the medium during growth resulted in up to 90% competence, while addition of retinyl acetate instead resulted in as low as 10% competence. Hydrocortisone partially antagonized the effect of retinyl acetate when both agents were added together. Involucrin levels, measured by radioimmunoassay, were modulated essentially in parallel with envelope competence under the various conditions tested. When the cells were grown in medium supplemented with hydrocortisone, the levels shortly after confluence were over 50-fold higher than in sparse cultures. Regardless of hydrocortisone or retinyl acetate addition, less than 1% of the cells were competent in sparse cultures of growing cells, but up to 90% exhibited this property after growth arrest in serum-free medium containing hydrocortisone. High levels of competence were correlated with cessation of cell division but not with loss of colony-forming efficiency; under optimal conditions, two-thirds of the cells were capable of both envelope formation and colony initiation. Normal human epidermal cells showed a 4- to 5-fold increase in envelope competence from sparse to confluent culture but were insensitive to the suppressive effect of retinyl acetate. The results suggest that some potential differentiated character of malignant keratinocytes may be suppressed in vivo by physiological agents such as vitamin A.

The destruction of rabbit articular cartilage after intra-articular injections of hydrocortisone acetate was investigated using histological, biochemical, and tracer methods. Fissures and cysts increased in number as increasing amounts of hydrocortisone were given. A linear decrease of hexosamine to less than 50 per cent after twelve injections was accompanied by insigificant changes in deoxyribonucleic acid and hydroxyproline content. The synthesis of proteoglycans and proteins was reduced to one-third, while the production of collagen dropped to less than one-fifth. The changes in thymidine incorporation were not significant. Based on these data, a model indicating the sequence of events which leads to joint destruction after intra-articular injections of glucocorticoid is proposed.