Suppression of lymphocyte proliferation and regulation of dendritic cell phenotype by soluble mediators from rat lacrimal epithelial cells.

Lacrimal epithelial cells appear to constitutively secrete autoantigens to their underling stroma. The present experiments address the hypothesis that they also secrete soluble factors that regulate immune responses. Epithelial cells, spleen cells and lymphocytes were obtained from rabbits or rats and cultured in various configurations. Monocytes from rat bone marrow were matured to dendritic cells (DC) ex vivo. Proliferation was measured by [3H]-thymidine incorporation; surface MHC Class II and CD86 using flow cytometry; and mRNA relative abundances using real time RT-PCR. Microporous culture inserts containing rat lacrimal cells inhibited proliferation of rabbit lymphocytes co-cultured with autologous lacrimal cells and of rat lymphocytes co-cultured with TNF-alpha-stimulated DC. They inhibited CD86 and MHC Class II surface expression by maturating DC and reversed surface expression of CD86 but not MHC Class II by partially matured DC. Subsequent exposure of partially matured DC to mediators from rat lacrimal cells reversed the ability to stimulate lymphocyte proliferation. TGF-beta(1) and IL-10 mRNAs increased somewhat when rat lacrimal cells were isolated but decreased markedly in rabbit lacrimal cells. Antibodies to TGF-beta prevented soluble factors from rat lacrimal cells from inhibiting proliferation of rabbit lymphocytes co-cultured with rabbit lacrimal cells, but recombinant TGF-beta alone did not mimic the soluble factors. IL-10 immunopositivity was detected in epithelial cells of interlobular ducts and occasional interstitial cells in rabbit lacrimal gland. Rat lacrimal epithelial cells secrete TGF-beta and other factors that synergize to suppress lymphocyte proliferation and regulate DC maturation. Interlobular duct epithelial cells in rabbit lacrimal glands may express similar functions.

Associations between imprinted gene differentially methylated regions, appetitive traits and body mass index in children.

BACKGROUND: Knowledge regarding genetic influences on eating behaviours is expanding; yet less is known regarding contributions of epigenetic variation to appetitive traits and body mass index (BMI) in children. OBJECTIVE: The purpose of this study was to explore relationships between methylation at differentially methylated regions (DMRs) of imprinted genes (insulin-like growth factor 2/H19 and Delta-like, Drosophila, homolog 1/maternally expressed gene 3) using DNA extracted from umbilical cord blood leucocytes, two genetically influenced appetitive traits (food responsiveness and satiety responsiveness) and BMI. METHODS: Data were obtained from participants (N = 317; mean age = 3.6 years; SD = 1.8 years) from the Newborn Epigenetic STudy. Conditional process models were implemented to investigate the associations between DMRs of imprinted genes and BMI, and test whether this association was mediated by appetitive traits and birthweight and moderated by sex. RESULTS: Appetitive traits and birthweight did not mediate the relationship between methylation at DMRs. Increased insulin-like growth factor 2 DMR methylation was associated with higher satiety responsiveness. Higher satiety responsiveness was associated with lower BMI. Associations between methylation at DMRs, appetitive traits and BMI differed by sex. CONCLUSIONS: This is one of the first studies to demonstrate associations between epigenetic variation established prior to birth with appetitive traits and BMI in children, providing support for the need to uncover genetic and epigenetic mechanisms for appetitive traits predisposing some individuals to obesity.

Changes in basic fibroblast growth factor coincident with estradiol-induced hyperplasia of the anterior pituitaries of Fischer 344 and Sprague-Dawley rats.

Adult female Fischer 344 (F344) and Sprague-Dawley (SD) rats were treated with estradiol via Silastic implants for 10 and 20 days. This treatment period in F344 rats is sufficient to produce dramatic hyperplasia of anterior pituitary lactotropes, activation of folliculo-stellate cells (FS) as phagocytes, and reorganization of the blood supply, i.e. hemorrhagic lakes and arteriogenesis from vessels in the adjacent meninges. Estradiol-treated SD rats do not demonstrate a comparable response. We now report intense focal concentrations of cells immunopositive for basic fibroblast growth factor (FGF) in estradiol-treated F344 rats predominantly near the posterolateral edge of the anterior pituitary, a zone rich in gonadotropes and lactotropes. Immunostaining for FGF, by both light and electron microscopy, revealed that the immunopositive cells were gonadotropes, and that the immunoprecipitate was cytosolic and was most abundant in the cytosol facing the capillaries. Immunostaining for extracellular matrix-associated FGF also revealed foci of positivity at the postero-lateral edge. Estradiol-treated SD rats did not reveal comparable localization for FGF. Morphological analysis and immunolocalization of S-100 protein, a marker for FS cells, revealed that the periphery of the anterior pituitary of estradiol-treated F344 rats included numerous disrupted gonadotropes and, furthermore, was largely devoid of FS cells. This zone was more intact in control F344 rats, but lacked FS cells. The peripheral parenchyma of control and estradiol-treated SD rats was intact compared to that of F344 rats and consistently included FS cells. These results suggest that disruptions of gonadotropes at the pituitary periphery may release FGF, which could then stimulate angiogenesis from blood vessels within the adjacent meninges. The resultant systemic blood supply would stimulate lactotrope hypertrophy and hyperplasia. Since FS cells are known phagocytes within the anterior pituitary, their absence from the periphery of F344 rats may intensify or prolong the effect of the peripherally released FGF.

Isolated growth hormone deficiency: immunocytochemistry.

Light and electron microscopic immunocytochemistry of the pituitary gland of a patient with documented isolated GH deficiency revealed parenchyma rich in GH positive cells. Immunoprecipitate was localized consistently over membrane-enclosed secretory granules whose features were characteristic of those in normal somatotrophs. No profiles of exocytosis of the contents of the secretory granules were observed. These observations demonstrate that the type I recessive form of isolated GH deficiency may be associated with apparently normal appearing somatotrophs which contain immunoreactive GH.

Morphological evidence for the presence of arteries in human prolactinomas.

This study was undertaken to clarify the vascular anatomy of human prolactinomas and specifically to determine whether arteries were present. Sixteen prolactinomas were studied by electron microscopy. The presurgical diagnosis of the tumors as prolactinomas was based on the findings of hyperprolactinemia and radiographic abnormalities, and was confirmed by the electron microscopic features of the specimens as well as by immunocytochemical staining for PRL. In addition to the presence of fenestrated endothelial cells, which are characteristic of the normal capillaries of the anterior pituitary, 13 of the 16 prolactinomas contained arteries. These arteries ranged from well formed vessels with multiple layers of smooth muscle cells to abnormal terminal arterioles, i.e. vessels with fenestrated endothelium surrounded by a variable number of smooth muscle cells. Arteries were not found in anterior pituitaries from 8 patients with no pituitary disease. In the prolactinomas, smooth muscle cells also were found, either isolated in the pericapillary connective tissue space or in small cords some distance from the vessel lumen. The results suggest that vascular changes, including arteriogenesis, occur in prolactinomas (and possibly other types of pituitary tumor). The arteries entering the anterior pituitary directly could be congenital or develop during formation of the tumor. An arterial blood supply to a region of the anterior pituitary could result in the escape of that area from hypothalamic regulation, since systemic blood contains negligible levels of hypothalamic hormones. In the case of PRL-secreting cells, which are tonically inhibited by the hypothalamic hormone dopamine, this would result in hypertrophy, hyperplasia, and possibly tumorigenesis.

Spontaneous tumors in aging female mice are more prevalent in the lateral pituitary zones.

The localization of gross and microscopic spontaneous pituitary tumors was examined in aging female C57BL/6J mice. These tumors were lactotroph adenomas, by morphological and immunocytochemical criteria. Each lobe of the pars distalis was divided into three zones of approximately equal size and the number of tumors in each zone was counted. Twelve out of 30 tumors were located entirely within the most lateral zone. An additional 14 tumors occurred in both the most lateral and the interjacent zones. Thus, almost 90% of the observed tumors were localized in more lateral zones of the pars distalis (Chi-squared test, p less than 0.01). These findings support a hypothesis that lower portal blood dopamine levels reaching lateral portions of the pars distalis are a factor in the higher incidence of lactotroph adenomas in these zones.

The development of mammotroph adenomas in pituitaries of aging female C57BL/6J mice.

The development of pituitary adenomas in female C57BL/6J mice was studied over the lifespan. Mammotrophs and somatotrophs, identified by immunocytochemical staining, appeared normal in 3-4 month old animals, with the exception of an occasional hypertrophic mammotroph and, in one gland, a nest of such cells. At 15 months, hypertrophic mammotrophs, somatotrophs and unstained cells occurred singly or in areas of hyperplasia; a mammotrophic adenoma was present in one gland. At 22 months, pituitaries contained mammotroph adenomas or adenomalike mammotroph clusters within hypertrophic, hyperplastic areas as seen in 15 month old glands. Pituitaries from all 28-30 month old mice contained mammotroph adenomas; nests of hypertrophic cells were present in 5 of 8 glands studied. These results suggest that mammotrophic adenomas may arise from hypertrophic mammotrophs, which are often found in nests with other hypertrophic pituitary cell types. However, this study does not exclude the possibility that mammotroph adenomas and hypertrophic mammotrophs occur independently.

Secure cranial-mount intrathecal cannulation - a spinal administration technique in the rat.

In order to identify an acute spinally mediated pharmacological effect of a bioactive substance, without incurring untoward supraspinal effects, it is necessary to administer the agent locally onto the spinal cord. The procedure delineated herein presents a modern technique to install a stable, permanent indwelling thecal cannulae with a cranially mounted aperture, and details a simple, repeatable administration system. These methods facilitate a quick, noninvasive spinal drug microadministration that is most useful for differentiation of the locus of pharmacological action without the behavioral disruption associated with other administration methodologies.

Genetic selection for nicotine activity in mice correlates with conditioned place preference.

Genetically heterogenous stock (HS) mice are being used to develop lines which have differential locomotor response to subcutaneously administered (0.75 mg/kg) nicotine. These groups of nicotine-depressed, nicotine-activated or randomly bred control mice were tested as to conditioned place preference using the same dose of nicotine employed to determine their locomotor performance in activity tests. Results indicate that the nicotine-activated mice showed a significantly greater preference to nicotine when compared to the nicotine-depressed mice; this effect was seen in the first generation and continued in the more recently tested third generation. Evidence is offered to support the hypothesis that it is the stimulatory effects of drugs (of abuse) that can be directly correlatable with the strength of their reinforcing effect upon behavior.

Anaphylaxis following the use of bacitracin ointment. Report of a case and review of the literature.

A 52-year-old woman suffered an acute anaphylactic reaction to topically applied bacitracin. An investigation disclosed the presence of concurrent type I and IV hypersensitivity to bacitracin. We review five other cases of anaphylactic reactions to topical medications and describe the clinical characteristics common to all six cases.

Cutaneous malignant lymphoma: a clinical and pathological study of 13 cases.

Thirteen cases of malignant lymphoma, exclusive of mycosis fungoides, with initial involvement of the skin, were selected from 391 consecutive patients with non-Hodgkin's lymphoma. Their clinical course was correlated to histological subtype. All patients (5/5) with 'favourable' histology had single indurated plaques without extracutaneous involvement, and remained in complete remission for at least 5 years after local radiation therapy. In contrast, all those with 'unfavourable' histology not lost to follow-up (7/7) suffered relapse or died within a 2-year period. They generally had multiple skin lesions and extracutaneous involvement at diagnosis or shortly afterwards. We conclude that histological subtype is an important variable in predicting clinical course in those with cutaneous malignant lymphoma.

Levonorgestrel concentrations during 7 years of continuous use of Jadelle contraceptive implants.

Serum levonorgestrel concentrations were assayed in a multicenter, 7-year study of 199 users of Jadelle rod implants. We examined drug levels, patterns of changes, factors affecting drug levels, and concentrations at which pregnancies occurred. Mean levonorgestrel concentrations declined from 435 pg/mL at 1 month of use to 64% of that value (280 pg/mL) at the end of 3 years. Between the end of the third and fifth years neither mean nor median serum levels varied markedly. At 5 years the mean concentration was again 64% of the first month's mean. Declining levels were observed thereafter through the end of 7 years when the mean, 224 pg/mL, was 52% of the 1-month value. Last measured drug concentrations of women who became pregnant during Jadelle use had mean and median values of 152 and 144 pg/mL, respectively, and a maximum value of 180 pg/mL. Analyses indicated ponderal index, body weight, duration of use, and a single clinical center were the most important variables affecting measured levonorgestrel levels. Approximately one-third of assays in the sixth and seventh years were found to be below 180 pg/mL, suggesting that Jadelle levonorgestrel implants would not maintain sufficiently high levels of effectiveness against pregnancy after 5 years and that heavier women would then be at greater risk of pregnancy.

Prolonged effectiveness of Norplant(R) capsule implants: a 7-year study.

Soft tubing Norplant(R) contraceptive implants were studied in 1210 women for 7 years to measure the duration of effectiveness and the magnitude of the pregnancy rates over that time. Mean age at enrollment was 27.4 years. Of the enrollees, 42% were US residents. One-sixth (16.1%) weighed >/=70 kg at the time of implant placement. At the end of 5 years, the cumulative pregnancy rate was 1.1/100; at the end of 7 years, it was 1.9/100. No pregnancies occurred to any of the 400 women who enrolled in the study at age >/=30 years and who weighed <100 kg. Among women aged 18-33 years, the 7-year Norplant pregnancy rates are comparable to the median pregnancy rates of tubal sterilization methods for women of the same age and duration of use. For women aged >/=34 years, without regard to weight at admission, the 7-year effectiveness of soft tubing Norplant equals or surpasses that of tubal sterilization. For continuing implant users, annual pregnancy rates <1.0/100 in years 6 and 7, together with low cumulative pregnancy rates, testify that Norplant capsule implants remain highly effective for 7 years.

Direct microinjection of cathinone into the rat brain produces discriminative stimuli.

Rats were trained to discriminate IP administration of 800 micrograms/kg cathinone using a food-motivated, two-lever discrimination procedure. Following training, 800 micrograms/kg cathinone discrimination was produced (generalized) by lower cathinone doses in a dose-responsive manner after IP administration; an ED50 value of 330 micrograms/kg was calculated. Subsequently, guide cannulae were implanted into the lateral ventricle and bilaterally into the nucleus accumbens. After recovery, injections were made via cannulae that extended 0.5 mm past the tip of the guide cannulae. ICV administration of 256 micrograms cathinone/rat produced discriminative responding on the cathinone-appropriate lever to the same degree as did the peripherally administered training dose of cathinone. Decreasing ICV doses produced decreased discriminative performance and allowed the calculation of an ED50 value of 90.5 micrograms. Likewise, administration of 64 micrograms cathinone/nucleus accumbens (for a total of 128 micrograms/rat) substituted for the IP training dose of cathinone. These results evidence the central mediation of the cathinone-induced discriminative stimulus cue and show that administration of cathinone into the nucleus accumbens is sufficient to produce these stimuli. Thus, these data suggest that receptors in the nucleus accumbens are important for the discrimination of this psychostimulant.

Role of radiation therapy in the management of cutaneous malignant melanoma.

Traditionally, cutaneous malignant melanoma is regarded as a radioresistant tumor. Recently, however, an increasing number of clinical studies have refuted this notion. The authors examined the role of radiation therapy in the palliative and/or adjuvant treatment of cutaneous malignant melanoma. The records of 69 patients with cutaneous malignant melanoma were reviewed. Twenty-five patients with extensive regional lymph node involvement received adjuvant radiation therapy after primary surgical treatment, and the remainder received palliative radiation therapy. The therapeutic significance of fraction size was analyzed. In the palliative radiation therapy group, the response rate was 52% with a fraction size < or = 300 cGy and 35% with a larger fraction size (p > 0.05, NS). Local regional control rates after adjuvant radiation therapy using conventional fractionation and larger fraction size were 87% and 82%, respectively (p > 0.05, NS). Radiation therapy is effective in the management of cutaneous malignant melanoma. It plays an important role in the palliation of metastatic disease and as an adjuvant treatment. No advantage in using a large fraction size over conventional dose schedules was found.

Application of Karasek's demand/control model a Canadian occupational setting including shift workers during a period of reorganization and downsizing.

PURPOSE: To apply Karasek's Job Content Model to an analysis of the relationships between job type and perceived stress and stress behaviors in a large company during a period of reorganization and downsizing. DESIGN: Cross-sectional mail-out, mail-back survey. SETTING: A large Canadian telephone/telecommunications company. SUBJECTS: Stratified random sample (stratified by job category) of 2200 out of 13,000 employees with a response rate of 48.8%. MEASURES: Responses to 25 of Karasek's core questions were utilized to define four job types: low-demand and high control = "relaxed"; high demand and high control = "active"; low demand and low control = "passive", and high demand and low control = "high strain." These job types were compared against self-reported stress levels, perceived general level of health, absenteeism, alcohol use, exercise level, and use of medications and drugs. Similar analyses were performed to assess the influence of shift work. RESULTS: Employees with "passive" or "high strain" job types reported higher levels of stress (trend test p < .0001); poorer health (trend test P = .006); and higher levels of absenteeism (trend test p < .0001). More shift workers reported themselves in poor or fair health (chi-square p = .018) and reported high levels of stress at home (chi-square p = .002) than nonshift workers. The relationships between job type and levels of stress, health and absenteeism, however, held for nonshift workers as well. CONCLUSIONS: Job types with high demand and low control were associated with increased stress, increased absenteeism, and poorer self-concept of health. The demand/control model of Karasek and Theorell was validated in this setting with respect to stress and some stress-associated attitudes and behaviors.

Is cellular disruption the mechanism of release of basic fibroblast growth factor from anterior pituitary gonadotropes?

Adult female Fischer 344 (F344) and Sprague-Dawley (SD) rats, intact and ovariectomized (10-30d), have been used for immunolocalization of basic fibroblast growth factor (FGF). Tissues were selected from three specific sites (postero-lateral, lateral wing, and anterior wedge) of the periphery of the anterior pituitary (AP) carefully maintaining the association between the gland and the highly vascular meningeal connective tissue which envelopes it. In both rat strains, most of the periphery of the AP was characterized by intact parenchymal cells delimited from the meningeal connective tissue by an intact basal lamina. However, foci also were evident in which parenchymal cells projected directly into the connective tissue without a basal lamina intervening. These zones, designated the Non-Delimited Peripheral Parenchyma (NDPP), were present minimally in control rats, but were more numerous in ovariectomized rats. Profiles of focally disrupted gonadotropes were evident within the NDPP of 20-30d ovariectomized rats juxtaposed against intact, granulated parenchymal cells. Partially disrupted gonadotropes also were evident within the peripheral parenchyma within approximately 100 mu of the edge, and occasionally the disruptions resulted in an association of neighboring gonadotropes as a syncytium. FGF was localized only within the cytosol of gonadotropes, i.e., cells immunopositive for LH-beta and FSH-beta subunits. Gonadotropes nearer to the edges of the AP, especially the postero-lateral edge, were the most intensely stained. Electron microscopy and immunostaining for S-100 protein, a marker for folliculo-stellate cells (FSC), demonstrated that in intact and ovariectomized SD rats FSC were present in all peripheral zones of the AP, whereas portions of the postero-lateral periphery of the AP of intact and ovariectomized F344 rats often lacked FSC. We propose that FGF may be released from the cytosol of gonadotropes by a mechanism of cellular disruption. FGF released at peripheral sites of the AP would be well-positioned to stimulate angiogenesis from systemic blood vessels within the meninges. Since FSC are known phagocytes within the AP, their consistent presence in the periphery of the AP of SD rats may help regulate the effects of the released FGF, and by contrast, their absence in F344 rats may intensify or prolong the effects of released FGF. Such differences may underlie the higher incidence of pituitary tumors in F344 rats.

Urolithiasis associated with hypercalciuria.

Fifty male patients with urolithiasis (UL), associated with idiopathic hypercalciuria (IH), were studied in comparison to a group of 18 male normocalcemic patients with inactive calcium stone disease of unknown etiology. In the group of IH-UL, in addition to hypercaliuria, statistically significant hyperphosphaturia with decreased tubular reabsorption of phosphate and hyperuricemia were observed; there was a tendency to hypophosphatemia although non-significant. In 36% of the IH-UL patients the first episode of renal colic appeared at age 40 to 50. Thirty-eight per cent of the IH-UL patients had recurrent stone formation. Twenty per cent of the IH-UL patients had a family history of urolithiasis. Forty-six per cent of all stones contained oxalate in addition to calcium, and 25% of the stones contained oxalate and phosphate.

A case of acute cerebral ischemia following inhalation of a synthetic cannabinoid.

OBJECTIVE: Synthetic cannabinoids are increasingly used in the United States as marijuana substitutes. However, reports of severe toxicity, resulting from their use, are limited. We present the case of acute cerebral infarction following synthetic cannabinoid inhalation. CASE REPORT: A 33-year-old man with no significant medical history presented at the emergency department with right-sided weakness and aphasia. He had smoked a synthetic cannabinoid (SC) product called "WTF" prior to the onset of symptoms. Physical examination showed right hemiparesis, dysarthria, and aphasia. Laboratory evaluation, electrocardiography, and computed tomography (CT) of the head were unremarkable. Following administration of intravenous tissue plasminogen activator, his symptoms improved. A repeat head CT showed acute infarction in the left insular cortex. His hypercoagulability panel was unremarkable, and the patient was discharged neurologically intact. Urine toxicology results were unremarkable. Analysis of the product by gas chromatography-mass spectrometry (GC-MS) procedure confirmed the presence of a synthetic cannabinoid known as XLR-11. CONCLUSION: XLR-11 has previously been associated with acute kidney injury in humans. However, there are no reports of it causing acute cerebral ischemic events. The close temporal association between XLR-11 inhalation and his stroke is concerning. Acute cerebral infarction may occur after XLR-11 use in healthy patients.

Systematic variations in immune response-related gene transcript abundance suggest new questions about environmental influences on lacrimal gland immunoregulation.

PURPOSE: Retrospective analyses were undertaken to assess the hypothesis that environmental variables influenced immunophysiological status of lacrimal glands from untreated female rabbits that had been housed out-of-doors until they were acquired for use as controls for experimental studies. MATERIALS AND METHODS: Rabbits were euthanized within 5 days of arrival at University Vivaria. Glands were divided for histology and RNA extraction. Transcript abundances were determined with real time RT-PCR. Sections were stained for CD18 and rabbit thymic lymphocyte antigen. Environmental variables assessed were mean daily high temperature, low humidity, high temperature/low humidity ratio, and days with above average temperature/humidity ratio ("adverse days") during the prior 30 days. RESULTS: Spearman's analyses revealed numerous significant correlations. Numbers of T cells and abundances of mRNAs for CD8; CCL2, and CCL4; IL-1α and IL-1ß; the T(H)1 cytokine, IL-2; and the T(H)2- and B cell cytokines, IL-4, IL-6, IL-10, APRIL, and BAFF, all increased with adverse days, while IFN-γ mRNA abundance decreased. Glands from the group exposed to the most adverse days remained free of immunopathological lesions. Glands from the group exposed to the highest temperatures fell above the regression curves for IL-4, APRIL, and BAFF calculated for the other groups and had significantly higher abundances of mRNAs for prolactin, IL-18, CCL21, CCL28, CXCL8, and CXCL13. One of six glands from this group contained small immune cell aggregates; the others appeared normal. The only gland that presented with frank histopathology was from a group that had experienced benign conditions. CONCLUSIONS: Increasing adverse days correlated with increasing abundances of transcripts, including mRNAs for IL-2, IL-10, and CD8, outside the T(H)1/T(H)2 paradigm. The findings raise intriguing questions as to whether and how such changes might be associated with homeostatic phenomena.