Enhanced hematopoiesis by hematopoietic progenitor cells lacking intracellular adaptor protein, Lnk.

Hematopoietic stem cells (HSCs) give rise to variety of hematopoietic cells via pluripotential progenitors and lineage-committed progenitors and are responsible for blood production throughout adult life. Amplification of HSCs or progenitors represents a potentially powerful approach to the treatment of various blood disorders and to applying gene therapy by bone marrow transplantation. Lnk is an adaptor protein regulating the production of B cells. Here we show that Lnk is also expressed in hematopoietic progenitors in bone marrow, and that in the absence of Lnk, the number and the hematopoietic ability of progenitors are significantly increased. Augmented growth signals through c-Kit partly contributed to the enhanced hematopoiesis by lnk-/- cells. Lnk was phosphorylated by and associated with c-Kit, and selectively inhibited c-Kit-mediated proliferation by attenuating phosphorylation of Gab2 and activation of mitogen-activated protein kinase cascade. These observations indicate that Lnk plays critical roles in the expansion and function of early hematopoietic progenitors, and provide useful clues for the amplification of hematopoietic progenitor cells.     

Preparation of PEG-conjugated fullerene containing Gd3+ ions for photodynamic therapy.

A novel photosensitizer with magnetic resonance imaging (MRI) activity was designed from fullerene (C(60)) for efficient photodynamic therapy (PDT) of tumor. After chemical conjugation of polyethylene glycol (PEG) to C(60) (C(60)-PEG), diethylenetriaminepentaacetic acid (DTPA) was subsequently introduced to the terminal group of PEG to prepare PEG-conjugated C(60) (C(60)-PEG-DTPA). The C(60)-PEG-DTPA was mixed with gadolinium acetate solution to obtain Gd(3+)-chelated C(60)-PEG (C(60)-PEG-Gd). Following intravenous injection of C(60)-PEG-Gd into tumor-bearing mice, the PDT anti-tumor effect and the MRI tumor imaging were evaluated. The similar O(2)(*-)generation was observed with or without Gd(3+) chelation upon light irradiation. Both of the C(60)-PEG-Gd and Magnevist(R) aqueous solutions exhibited a similar MRI activity. When intravenously injected into tumor-bearing mice, the C(60)-PEG-Gd maintained an enhanced MRI signal at the tumor tissue for a longer time period than Magnevist(R). Injection of C(60)-PEG-Gd plus light irradiation showed significant tumor PDT effect although the effect depended on the timing of light irradiation. The PDT efficacy of C(60)-PEG-Gd was observed at the time when the tumor accumulation was detected by the enhanced intensity of MRI signal. This therapeutic and diagnostic hybrid system is a promising tool to enhance the PDT efficacy for tumor.     

Effects of antithyroid drug therapy on blood glucose, serum insulin, and insulin binding to red blood cells in hyperthyroid patients of different ages

The mechanism of glucose intolerance in thyrotoxicosis was investigated in 119 patients with Graves's disease with careful consideration of the age-related deterioration of glucose tolerance. Before and after treatment of thyrotoxicosis with antithyroid drug, changes of blood glucose (BG) and serum immunoreactive insulin (IRI) in response to 50 g oral glucose tolerance test (OGTT) and insulin binding to red blood cell (RBC) were evaluated. In control subjects, the sigma IRI/sigma BG ratio after 50-g OGTT decreased progressively with age without significant change in absolute sigma IRI value, suggesting the occurrence of age-related insulin resistance. Glucose intolerance was much more apparent in hyperthyroid patients because of age-related relative decrease of insulin secretion. Such a decrease of insulin secretion was not found in age-matched postgastrectomy patients with a similar degree of hyperglycemia, however. Maximal binding of labeled insulin and number of insulin receptors of RBC were decreased in old patients but binding affinity was unchanged. Elevation of BG was partially suppressed when serum thyroxine (T4) and triiodothyronine (T3) were reduced to moderately supernormal levels, whereas sigma BG, sigma IRI, sigma IRI/sigma BG ratio, and insulin binding to RBC were all returned to normal when normal serum thyroid hormone concentration was maintained. Our data indicate that insufficient insulin secretion and reduced insulin action at the target cell are responsible, at least in large part, for age-related glucose intolerance in hyperthyroid patients.     

The relationship between cholelithiasis and diabetes mellitus: discussion of age, obesity, hyperlipidemia and neuropathy

We investigated the patients who underwent operation for cholelithiasis and the diabetic patients at our clinic in order to determine whether there was a significant relationship between the occurrence of cholesterol gallbladder stone and age, obesity, hyperlipidemia, diabetes mellitus and neuropathy. In 647 patients undergoing surgery, cholesterol gallstones were not highly associated with diabetes mellitus or hyperlipidemia, compared with calcium bilirubinate and black stones. Eighty-seven percent of the male operated patients and 88% of the female patients were over 40 years old of age. Of the female patients in whom gallstones were detected at surgery, 36% were obese. We found cholesterol gallbladder stone in 11.5% (males 11%, females 12%) of 208 diabetic patients at our clinic. All of them were over 40 years old. The prevalence of cholesterol gallbladder stones was related to the decrease in motor nerve conduction velocity in the male diabetic patients (p less than 0.05). We observed that method of treatment had no definite effect on the prevalence of gallbladder stones. Fifty-four percent of the diabetic patients was normolipidemic in both sexes. Obesity was present in 64% of the female cholesterol gallbladder stone patients. Our data suggest that age, obesity and poor contraction of the gallbladder could be high risk factors for cholesterol gallstone formation.     

The stimulatory effect of PACAP 38 on amylase release in dispersed rat pancreatic acini.

Pituitary adenylate cyclase activating polypeptide 38 (PACAP 38), a novel peptide of the vasoactive intestinal polypeptide (VIP) family, was shown to stimulate enzyme secretion in the dispersed rat pancreatic acini. The dose-response of pancreatic enzyme secretion to PACAP 38 was nearly identical with that to VIP. In the presence of a submaximal dose of PACAP 38 (1 nM), amylase release stimulated by an agonist working via the elevation of intracellular cyclic AMP (VIP, dibutyryl cAMP) was additionally responded, but the amylase release stimulated by an agonist via the elevation of cytosolic free calcium (carbachol, cholecystokinin) was potentiated synergistically. The present data suggest that PACAP 38 is a new candidate for the cAMP-mediated stimulant of pancreatic exocrine secretion.     

Evidence for Stimulation of Renal Gluconeogenesis by Catecholamines

Isoproterenol and norepinephrine (10(-4) M) significantly increased cyclic AMP formation and glucose production by the isolated tubules of the renal cortex of the rat. These effects were abolished by propranolol. Theophylline diminished the effects of the catecholamines on gluconeogenesis despite a marked augmentation in cyclic AMP concentration. In the absence of calcium ion in the incubation medium, isoproterenol stimulates cyclic AMP production, but has no effect on gluconeogenesis. It is concluded that catecholamines enhance gluconeogenesis in renal cortical tubules by the stimulation of beta adrenergic receptors. This effect is probably mediated through adenyl cyclase-cyclic AMP system and requires an adequate level of ATP and the presence of calcium ion.     

Diagnostic and epidemiological implications of regional differences in serum concentrations of proteins observed in six Asian cities.

The IFCC Committee on Plasma Proteins has been investigating regional differences for commonly assayed plasma proteins to determine whether universal reference intervals can be applied. As a part of this study, we launched an Asian project analyzing the concentrations of 13 serum proteins whose values are standardized to CRM470, and five newer analytes: retinol-binding protein (RBP), cystatin C (CysC), light-chain-kappa (L-kappa), and light-chain-lambda (L-lambda). In Tokyo, Seoul, Kuala Lumpur, Hong Kong, Taipei and Shanghai, serum samples were collected from 146 to 415 apparently healthy individuals with nearly equal gender ratios. All assays were performed in Tokyo on a Behring Nephelometer II (BN II). Seven chemical analytes (aspartate aminotransferase (AST), alanine aminotransferase (ALT), gamma-glutamyltransferase (gammaGT), creatinine, total cholesterol (TC), triglycerides (TG) and high-density lipoprotein cholesterol (HDL-C)) were also measured. These results were used for excluding individuals with possible latent clinical disorders. Positive acute phase reactants were consistently lower, and negative ones were higher, in Tokyo than those in other cities. The most conspicuous difference was observed in C-reactive protein (CRP). There were no regional differences in transferrin, albumin, or CysC. Creatinine was much lower in Tokyo despite comparable CysC levels. ALT and gammaGT were higher in Shanghai, Taipei and Seoul; gammaGT and TG were higher in Shanghai; and HDL-C was higher in Tokyo. Gender-related differences in reference intervals were observed for immunoglobulin (Ig)M, haptoglobin, RBP, transferrin, alpha2-macroglobulin (A2M), transthyretin, alpha1-acid glycoprotein, CysC, and C4 in all cities. Slight age-related differences were observed, irrespective of the region, in IgA and ceruloplasmin (increase) and A2M (decrease). Environmental factors and lifestyle seem to have a great influence on many commonly measured analytes.     

Appropriate radial clearance of ceramic-on-ceramic total hip prostheses to realize squeeze-film lubrication

OBJECTIVE: Estimation of appropriate radial clearance of ceramic-on-ceramic total hip prostheses to realize squeeze-film lubrication. BACKGROUND: Some clinical results show that severe wear occurs at ceramic-on-ceramic interfaces of total hip prostheses if the design parameters are improper. Appropriate design is required to realize the optimum lubrication of joint prostheses. DESIGN: Squeeze-film thickness of total hip prostheses with different radial clearances was numerically estimated under the physiological conditions of normal walking. METHODS: The changes in the fluid film thickness between ceramic balls and a ceramic cups with radial clearances of various values from 10 to 80 microm were numerically estimated under the assumption of normal walking conditions by means of elastohydrodynamic squeeze-film lubrication theory including the three-dimensional theory of elasticity. RESULTS: The minimum film thickness remained over 0.02 microm during a few walking steps with a radial clearance under 20 microm and a head diameter of 28 mm. On the other hand, the fluid film was squeezed out during a few steps when the radial clearance was greater than 30 microm. CONCLUSIONS: Squeeze-film lubrication can be realized in a ceramic-on-ceramic total hip prosthesis if the radial clearance is smaller than 20 microm on a head of 28 mm diameter. RELEVANCE: Fluid film lubrication is necessary to prevent the severe wear of ceramic joint prostheses. The present investigation shows which radial clearance of ceramic-on-ceramic hip prostheses is appropriate for squeeze-film lubrication under physiological conditions in daily action.     

Competitive repopulation assay of two gene-marked cord blood units in NOD/SCID/gammac(null) mice.

In multiunit cord blood transplantation, hematopoietic stem cells from each unrelated cord blood (UCB) unit competitively reconstitute the hematopoietic system in a recipient. To evaluate the fate of the progeny of each UCB unit and to determine the effects of graft-versus-graft reaction, we established a novel competitive repopulation assay using NOD/SCID/gammac(null) mice in which human T lymphocytes develop from CD34+ cells. CD34+ cells from each UCB unit were labeled with recombinant lentivirus vectors carrying genes encoding either enhanced green fluorescent protein (EGFP) or enhanced yellow fluorescent protein (EYFP). Hematopoietic chimerism composed of both EGFP+ and EYFP+ cells was stably maintained up to 6 months after transplantation with purified CD34+ cells; the ratio of EGFP+ to EYFP+ cells in peripheral blood and bone marrow posttransplantation was equivalent to the ratio of these cells at transplantation. However, when mononuclear cells from two UCB units were cotransplanted with CD34+ cells, engraftment was highly competitive, with cells from only one or the other of the two UCB units surviving. Further subfractionations of mononuclear cells indicate that the skewed chimerism that is often observed in clinical multiunit cord blood transplantation may be mediated by the cooperation of both CD4+ and CD8+ T cells. The assay established here will be a useful tool for analyzing hematopoietic reconstitution in clinical multiunit cord blood transplantation.