Estimation of absorption enhancement by medium-chain fatty acids in rat large intestine.

The absorption enhancement by the sodium salts of several fatty acids was investigated in rat large intestine for model compounds having a wide range of molecular weight. Sodium caprylate (C8), sodium caprate (C10), sodium laurate (C12), which are categorized in medium-chain fatty acid, and sodium oleate (C18:1), long-chain unsaturated fatty acid, were employed as lipoidal adjuvants. Phenol red (MW=354.4), glycyrrhizin (822.9), fluorescein isothiocyanate-dextran-4 (FD-4, 4400), FD-10 (9400) and FD-40 (38900) were selected as model compounds for the assessment of the enhancing effect of the lipoidal adjuvants. The absorption of phenol red was promoted at the highest level, about 20 times higher by C12 and C18:1 than the control. The absorption rate - time profiles calculated by deconvolution method showed that C12 takes effect most rapidly and efficiently. In the case of glycyrrhizin, four adjuvants including C12 showed almost the same improvement of the absorption, about 30-40 times larger than the control. C8 and sodium citrate did not significantly enhance the absorption of those model compounds. For FD-4, FD-10 and FD-40, C10, C12 and C18:1 revealed almost the same enhancing effect and the absorption of FD-4, FD-10 and FD-40 was enhanced about 80 times, 1000-1800 times and about 200 times, respectively, larger than the control. The enhancement ratio, the ratio of AUC with adjuvant to AUC of control, suggests that these lipoidal adjuvants would improve most efficiently the absorption of the compound having the molecular weight of around 10000. Furthermore, C12 was suggested to be an effective adjuvant for the compounds with the wide range of molecular weight.

Leptin and the pituitary.

In 1994, Zhang et al. of Rockefeller University in New York reported the first successful complementary DNA (cDNA) cloning of leptin by the positional cloning method. Leptin was identified as the gene of ob/ob mouse in genetic obesity syndromes. It has very strong food intake control, and body weight and energy expenditure. The name "leptin" derived from the Greek word leptos, meaning "thin." We hereby review major advances leading to our current finding of leptin, leptin receptor and its structure, the outline of homozygote, and also influence of leptin in the pituitary. (The structure of leptin) The mouse obese gene has been localized to chromosome 6. With human leptin gene on chromosome 7q31.3, its DNA has more than 15000 base pairs and consists of three exons and two introns. For bioactivation of leptin the importance of disulfide-binding site is suggested. Human leptin which replaced the 128-th arginine with glutamine has the function of an aldosteron antagonist, which is reported to have the function of athrocytosis inhibition. The resemblance of leptin precursor of human, mouse and rat is very high, i.e., mouse and rat homology is 96% and mouse and human homology is 83%. (The structure of leptin receptor) The mutant gene, which is the cause of obesity, was shown on map on diabetic mouse (db/db) chromosome 4, and it was proven to be the same as the leptin receptor gene cloned by Tartaglia et all. Further studies have found the Zucker fatty rat (fa/fa) to be incorporated into a linkage map of rat chromosome 5, whose region of rat is the equivalent to the region of conserved synteny of the db/db mouse gene. The leptin receptor is glycoprotein consisting of a single transmembrane-spanning component. The primary structure of leptin receptor belongs to the cytokine-class1 family, the single membrane-spanning receptor, and is highly related to the gp130 signal-transducing component of the interleukin-6 (IL-6) receptor, the granulocyte colony-stimulating factor (G-CSF) receptor, and the leukemia inhibitory factor (LIF) receptor. The leptin receptor is known to have at least six existing isoforms (Ob-Ra, b, c, d, e, f) from the difference in splicing. (Homozygote Mutation of Leptin and Leptin Receptor :Hormone Secretion Disorders) The point mutation of ob/ob mouse and the splicing mutation of db/db mouse show remarkable obesity and hyperphagia. These obesity models show a reproduction disorder with both the male and the female, and they develop with homozygote. The cause is thought to be the gonadotropin secretory abnormality in pituitary. Three family lines report the cases of this deficiency, and it is considered that the secretory abnormality in pituitary develops into hypogonadotropic. These patients show low value in plasma FSHbeta (follicle stimulating hormone-beta and LHbeta (luteinizing hormone-beta which are produced from pituitary, and the plasma GnRH (gonadotropin releasing hormone) level is also low. Furthermore, the leptin receptor deficient family line was reported in 1998, in which case only the homozygote developed. The plasma leptin concentration of normal human is about 8.0 ng/ml, and this case with leptin receptor deficiency has high value of 500-700 ng/ml, which is the equivalent to the db/db mouse. (Role of Leptin in Hypothalamus-Pituitary-Periphery Function) The role of leptin which regulates pituitary hormones suggests the promotion the GHRH (growth hormone releasing hormone) secretion in hypothalamus-pituitary axis, with the possibility of the rise in secretion of GH (growth hormone) in pituitary, i.e. effects of icv (intracerebroventricular) infusion of leptin has spontaneously stimulated GHRH, which promotes GH secretion in the normal rats. On the other hand, topical treatment of GH3 (derived from a rat pituitary GH-secreting cell line) with leptin directly inhibits cell proliferation. The obesity model animals (ob/ob, db/db, fa/fa) have equally plump body compared to the normal models, which shows signs of sufficient growth. (Localization and Functional Relevance of Leptin and Leptin Receptor in Rodents Pituitary) Aside from being the food intake inhibitor and the energy control factor, leptin takes part in controlling the pituitary hormones. Promoting the secretion of GH, PRL (prolactin), TSHbeta (thyroid stimulating hormone-beta, FSHbeta/LHbeta, and inhibiting the secretion of ACTH (adrenocorticotropic hormone) are the major changes of pituitary hormones which are brought on by leptin. The expressive localization is specific, and immunohistochemistry (IHC) method recognized leptin in granular state in FSHbeta, LHbeta and TSHbeta positive cells. In our biochemical examination, the bulk of the expression of leptin is recognized in fraction of the secretory granule. In particular, FSHbeta cells had the highest percentage rate of colocalized leptin in rat pituitary. On the other hand, leptin receptor has been reported to be found only in normal rat pituitary, human pituitary adenoma, and respective cell lines in pituitaries by the RT-PCR method until now, but we disclosed for the first time the localization of leptin receptor on the plasma membrane of GH-secreting cells with the IHC method that has not been cleared so far. These findings show that leptin and leptin receptor have been expressed in different cells, and that the rat pituitary glands entertain paracrine mechanism between leptin (FSHbeta/LHbeta cells) and leptin receptor (GH cells). The function of paracrine in this pituitary suggests a new point of view in hypothalamus-pituitary axis, and it shall be concerned with many aspects such as hormone secretions and proliferation/inhibition. (Human Pituitary Adenoma) Preliminary report of leptin and leptin-receptor relationship with pituitary adenoma that has secretion abnormality has been filed, and its manifestation is being observed by the RT-PCR. Leptin and leptin receptor are expressed in most adenoma, and it is thought to function by autocrine and paracrine pathway in the adenomas. Leptin has been located in ACTH-secreting adenoma most frequently, especially in ACTH carcinoma. The leptin receptor is detected in all adenomas with high percentage rate, with both long and short forms, and then many cases of nonfunctioning pituitary adenomas, compared with other adenomas, have been reported to be positive with both long and short forms of leptin receptor as detected by RT-PCR. The HP75 cell line is derived from the nonfunctioning pituitary adenoma, which produces FSHbeta and LHbeta. The expression of leptin receptor in nonfunctioning pituitary adenoma, and the suppression of HP75 multiplication may lead to the possible hypothesis of leptin becoming one factor for the treatment of pituitary adenoma, especially in gonadotropin adenomas.

Regional distribution of immunoreactive prolactin-releasing peptide in the human brain.

Regional distribution of prolactin-releasing peptide (PrRP) in the human brain was studied by radioimmunoassay. The antiserum raised against human PrRP-31 in a rabbit was used in the assay, which showed 100% cross reaction with PrRP-20 and no significant cross reaction with other peptides. The highest concentrations of immunoreactive-PrRP were found in hypothalamus (912 +/- 519 fmol/g wet weight, n = 6, mean +/- SEM), followed by medulla oblongata (496 +/- 136 fmol/g wet weight) and thalamus (307 +/- 117 fmol/g wet weight). On the other hand, immunoreactive-PrRP was not detected in frontal lobe or temporal lobe (<50 fmol/g wet weight). Sephadex G50 column chromatography of the immunoreactive-PrRP in the hypothalamus and medulla oblongata showed three immunoreactive peaks; one peak eluting in the position of PrRP-20, one eluting in the position of PrRP-31 and one eluting earlier. Reverse phase high-performance liquid chromatography (HPLC) of these brain tissue extracts showed a peak eluting in the position of PrRP-20 and PrRP-31. The present study has shown for the first time the presence of immunoreactive-PrRP in the human brain. The immunoreactive-PrRP levels in the human hypothalamus were, however, lower than the levels of other neuropeptides with prolactin-releasing activity, such as thyrotropin-releasing hormone and vasoactive intestinal polypeptide.

Molecular cloning of rat p38 mitogen-activated protein kinase and it's osmotic regulation in rat kidney.

Rat p38 mitogen-activated protein (MAP) kinase cDNA was isolated from rat kidney cDNA library using a PCR cloning strategy. The deduced amino acid sequence consists of 360 amino acids and shares 95.3% similarity with human p38 MAP kinase. The message for rat p38 MAP kinase was about 3.4 kilobases and was highly expressed in the kidney. In water-deprived rat kidneys, the steady-state levels of p38 MAP kinase mRNA increased about 2.7-fold as compared with those of control rats. This result suggests that p38 MAP kinase may play an important role in the osmoregulation in the kidney.

Laryngotracheal separation under local anaesthesia for intractable salivary aspiration: cricoidectomy with fibrin glue support.

Chronic salivary aspiration can cause life-threatening pneumonia in a patient whose laryngeal function is completely lost. We report a patient who had laryngotracheal separation with cricoidectomy under local anaesthesia. The simplicity and reliability of the procedure were improved by using fibrin glue and the outcome was most satisfactory.

Immunocytochemical localization of adrenomedullin-like immunoreactivity in the human hypothalamus and the adrenal gland.

Adrenomedullin is a potent vasodilator peptide that was isolated from pheochromocytoma. Localization of adrenomedullin-like immunoreactivity was studied by immunocytochemistry in the human hypothalamus and adrenal gland. Adrenomedullin-immunoreactive cell bodies were found in the paraventricular, supraoptic and infundibular nuclei of the hypothalamus. Both magnocellular and parvocellular cells of the paraventricular nucleus were positively immunostained. Adrenomedullin-like immunoreactivity was localized in the adrenal medulla. No positive immunostaining was observed in the vascular endothelium, vascular smooth muscle cell or adrenal cortex. The preabsorption of the antiserum with synthetic human adrenomedullin (1-52) abolished the immunostaining. These findings indicate that adrenomedullin-like immunoreactivity is localized in the paraventricular, supraoptic and infundibular nuclei as well as in the adrenal medulla, and suggest that adrenomedullin acts as a neurotransmitter, a neuromodulator or a neurohormone in the human hypothalamus.

Melanin-concentrating hormone in human and rat.

Melanin-concentrating hormone (MCH) is a neuropeptide originally isolated from chum salmon pituitaries. To explore physiological roles of MCH in mammals, we studied the regional distribution of immunoreactive MCH in the rat tissues and the presence of immunoreactive MCH in human adrenal glands, adrenal tumors and plasma by radioimmunoassay, and the expression of MCH mRNA in rat tissues and human brain tissues by Northern blot analysis. Immunoreactive MCH was present in every region of rat brain and neurointermediate lobe of the pituitary gland, with the highest concentrations found in the hypothalamus (48.3 +/- 6.6 pmol/g wet weight, mean +/- SEM, n = 6). The immunoreactive MCH in rat hypothalamus, frontal lobe, and pons and medulla oblongata was eluted in the position of synthetic human/rat MCH in reverse-phase high-performance liquid chromatography. No immunoreactive MCH was detected in the rat peripheral tissues. Northern blot analysis showed that a single species of MCH mRNA (approximately 1 kb) was expressed specifically in the rat and human hypothalamus, but not detectable in other regions of brain or rat peripheral tissues. Immunoreactive MCH was not detected in human adrenal glands (< 0.5 pmol/g wet weight, n = 9) or adrenal tumors including pheochromocytomas. Immunoreactive MCH were not detected in plasma obtained from human healthy subjects (< 0.25 pmol/l) and rat (< 0.25 pmol/l).(ABSTRACT TRUNCATED AT 250 WORDS)

A substrain of NZB mouse as an animal model of autoimmune inner ear disease.

A substrain of an autoimmune-prone mouse, NZB/kl, was found to show spontaneous elevation of the auditory brainstem response (ABR) threshold with age. Morphological examination of the inner ear in NZB/kl mice with high ABR thresholds revealed pathological changes confined to the stria vascularis, including marked thickening of the capillary basement membrane which contained many foamy structures, and vacuolar degeneration of the intermediate cells. Circular or granular IgM deposits and some IgG deposits were found in the stria vascularis in the mice with high ABR thresholds, suggesting that deposits of immune complexes (mainly IgM antibodies) could cause strial damage that resulted in the ABR threshold elevation. Another substrain of NZB mice, NZB/san, showed lower levels of IgM immune complexes and anti-ss DNA antibodies, and did not develop either inner ear morphological changes or a high ABR threshold. NZB/kl mice may provide a useful animal model for studying the mechanism of autoimmune inner ear disease.

Pituitary adenylate cyclase activating polypeptide (PACAP)-like immunoreactivity in human hypothalamus: co-localization with arginine vasopressin.

Pituitary adenylate cyclase activating polypeptide (PACAP) is a novel hypothalamic peptide consisting of 38 amino acids (PACAP1-38) with a potent stimulatory action on adenylate-cyclase in rat pituitary. The presence of PACAP-like immunoreactivity in human brain was studied by radioimmunoassay. Co-localization of PACAP with arginine vasopressin and oxytocin was investigated by immunocytochemistry in the human hypothalamus. Immunoreactive PACAP was detected in all regions of human brain (cortex, thalamus, hypothalamus, pons and hemisphere of cerebellum) with the highest levels found in the hypothalamus (8.5 +/- 1.9 pmol/g wet weight, n = 4, mean +/- S.E.M.). High performance liquid chromatography of the human hypothalamic extract showed that approximately 50% of the immunoreactive PACAP was eluted in the position of PACAP1-38. Immunocytochemical studies showed the presence of PACAP immunoreactive neurons in the paraventricular and supraoptic nuclei of human hypothalamus. PACAP co-localized with arginine vasopressin in magnocellular cells of these nuclei. These findings suggest that PACAP1-38 plays important physiological roles in the human hypothalamus.

Calcitonin gene-related peptide in the human hypothalamus.

Calcitonin gene-related peptide (CGRP)-like immunoreactivity (LI) in the human hypothalamus was investigated by radioimmunoassay and by immunocytochemistry. CGRP-LI was detected from two hypothalami obtained at autopsy (2.1 and 7.0 ng/g wet tissue) by radioimmunoassay. Reverse phase high performance liquid chromatography revealed that most of the CGRP-LI in the human hypothalamus was eluted in an identical position with synthetic human CGRP. For immunocytochemistry, human hypothalami obtained at autopsy were fixed and cryostat-sectioned at 40 microns. Free floating sections were immunostained with antibody to CGRP. CGRP-immunoreactive cell bodies were found in the supraoptic nucleus, paraventricular nucleus and infundibular nucleus. These findings indicate that CGRP exists in the cell bodies of the supraoptic nucleus, paraventricular nucleus and infundibular nucleus in the human hypothalamus and CGRP may play some roles in the endocrine and other functions of the human hypothalamus.