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Publications and source records attributed to L Hu.
Allele-specific MVR-PCR can be performed on genomic DNA to sample the diversity of allelic structure at MS31A (D7S21 locus). Three polymorphic positions in MS31A 5' flanking DNA were typed from 100 unrelated Chinese in this study. The results showed that single alleles could be mapped from genomic DNA in above 80% samples of Chinese in this locus. The distribution of haplotypes of unrelated Chinese was also reported in this paper.
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A previous study in conscious dogs showed that the normal hypertensive response to short-term nitric oxide synthesis inhibition was markedly attenuated during angiotensin II-AT1 receptor inhibition. However, whether angiotensin plays an important cardiovascular role in the dog during long-term nitric oxide synthesis inhibition has not been determined and was therefore the goal of this investigation. Studies were conducted in 16 conscious dogs that received angiotensin AT1 receptor inhibition with L158809 (N = 8) or vehicle (N = 8) for 12 d. During the last 6 d of this infusion, nitric oxide synthesis was inhibited by infusing NG-nitro-L-arginine methyl ester intravenously at 37.1 nmol/kg/min. In both the AT1 and vehicle groups, nitroarginine infusion significantly decreased the acetylcholine depressor response, glomerular filtration rate, renal plasma flow, and heart rate, and increased arterial pressure and renal vascular resistance in a similar manner, whereas it caused little change in the urinary excretion of sodium and water or in plasma renin activity. In conclusion, the long-term responses of arterial pressure, renal hemodynamics, and the renal excretion of sodium and water to nitric oxide synthesis inhibition were not significantly influenced by blockade of angiotensin AT1 receptors with L158809 in the dog.
OBJECTIVE: To investigate whether plasma angiotensin II (Ang II) determines the effects of the renin-angiotensin system or whether tissue uptake of renin and localized production of Ang II might account for any cardiovascular, renal, hormonal or drinking effect of circulating renin. DESIGN: Intravenous infusions of renin (0.6 ng/min; n = 10) and Ang II (3.5 ng/min; n = 10) that produce similar plasma Ang II levels were compared for 2 weeks with vehicle (n = 7) in conscious rats after a 1-week control period. Mean arterial pressure (MAP) and the heart rate were measured continuously. Hormones and renal function were measured twice weekly. Plasma Ang II and recovery data were measured in seven additional rats. RESULTS: In renin- and Ang II-infused rats, respectively, plasma Ang II increased similarly from 4.5 +/- 0.8 and 4.4 +/- 0.9 to 10.8 +/- 0.7 and 10.6 +/- 0.7 pg/ml and declined similarly in the second week to 7.0 +/- 1.1 and 7.0 +/- 1.5 pg/ml. Plasma renin increased from 4.2 +/- 0.7 to 21.7 +/- 1.3 and fell from 5.9 +/- 0.5 to 0.6 +/- 0.2 ng/ml per h respectively. Plasma prorenin fell similarly (> 70%); angiotensinogen was unchanged. MAP rose initially by 25.6 +/- 1.2 and 23.3 +/- 0.9 mmHg and by an additional 21.1 +/- 2.4 and 27.4 +/- 1.8 mmHg on days 5-8. The heart rate fell gradually but transiently by -11% in both. Although the initial MAP rise was slower in renin-infused rats (P< 0.05) MAP returned to baseline within 2 h after both infusions were stopped. Changes in renal vascular resistance, renal blood flow, glomerular filtration rate, urinary sodium, potassium and water excretion and water intake were not significantly different between renin- and Ang II-infused rats. CONCLUSIONS: Intravenous infusions of low doses of renin or Ang II into conscious rats increase MAP identically. MAP increases in two phases 5-8 days apart, in coordination with transient falls in the heart rate. Renin- and Ang II-induced chronic hypertension are identically sustained by very small increases in plasma Ang II. Blood pressure increases more slowly with renin infusions, consistent with tissue binding. Notwithstanding, no evidence was obtained for a physiological role of tissue-bound renin in causing the cardiovascular, renal, hormonal and drinking responses measured in this study.
Administration of high doses of glutamate (Glu) leads to selective neurodegeneration in discrete brain regions near circumventriclular organs of the early postnatal mouse. The arcuate nucleus-median eminence complex (ARC-ME) appears to be the most Glu-sensitive of these brain regions, perhaps because of the intimate relationships between its neurons and specialized astroglial tanycytes. To investigate the mechanism of Glu-induced neuronal loss, we administered graded doses of the sodium salt of glutamate (MSG) to postnatal mice, measured their plasma Glu concentrations, and performed microscopic analyses of the ARC-ME region 5 h after treatment. Nursing, 7-day-old mouse pups (CD1, Charles River, Hollister, Calif.) were injected subcutaneously with single doses of 0.1-0.5 or 1.0-4.0 mg of MSG per g BW, or with water vehicle alone. Mice were decapitated 5 h later and the brains immediately fixed by immersion in buffered aldehydes. Frontal vibratome tissue sections at comparable levels of the ARC-ME were examined by light microscopy. A dose of 4.0 mg MSG/g BW caused neurodegeneration throughout the ARC region, while 1.0 mg/g MSG resulted in less extensive damage. Injection of 0.2 mg MSG/g BW, which raised plasma Glu concentrations 17-fold after 15 min, was the minimum dose tested at which nuclear and cytoplasmic changes were observed in a small group of subependymal neurons near the lateral recesses of the third ventricle. Higher doses of 0.3-0.5 mg MSG caused injury to additional neurons situated farther laterally, but damage remained confined to the ventral region of the ARC nucleus. Ultrastructural examination showed some subependymal neurons with pyknotic nuclei, reduced cytoplasmic volume, and swollen subcellular organelles, while others had fragmented and condensed nuclear material. Immunostaining for tyrosine hydroxylase indicated that dopamine neurons were spared at the threshold dose, but suffered damage after higher doses of MSG. Immunostaining for Glu receptor subtypes revealed that 0.2 mg MSG/g BW enhanced neuronal expression of NMDAR1 and of GluR2/4, and that higher doses of MSG preferentially increased NMDAR1 expression in injured neurons. These results extend previous reports of Glu sensitivity in the ARC-ME region of 7-day postnatal mice. A dose of 0.2 mg MSG/g BW s.c. causes clear but discrete injury to specific subependymal neurons of undetermined phenotype near the base of the third ventricle. Slightly higher doses of MSG evoke damage of additional neurons confined to the ventral region of the ARC traversed by tanycytes. These same greater amounts of MSG promote dose-related increase in the expression of NMDAR1 more than of GluR2/4 in injured ARC neurons, suggesting that elevated Glu receptor levels may contribute to or be related to neuronal cell death. Taken together with previous findings, the data suggest that Glu responsitivity in the ARC-ME of the postnatal mouse may result from transient developmental conditions involving the numerical ratios and juxtaposition between tanycytes and neurons, expression of Glu receptors, and perhaps other ontogenetic factors which may not persist in the mature adult.
To create physiological models of the human renin-angiotensin system in transgenic animals, the component genes should be expressed in the correct tissues and cells and respond appropriately to physiological stimuli. We recently showed that mice carrying a 45-kb human renin genomic fragment, containing approximately 25 kb 5'-flanking DNA and 6 kb 3'-flanking DNA, express the transgene in a highly cell- and tissue-specific pattern. More importantly, in contrast to previous models, human renin in the circulating plasma of these mice is derived exclusively from the kidneys. In the present study, we tested the responses of both human and mouse renal renin expression and secretion of the 45-kb hREN transgenic mice to a variety of physiological and pharmacological stimuli. A sodium-deficient diet, angiotensin-converting enzyme inhibition, and beta1-adrenergic stimulation each increased both human and mouse plasma renin concentration significantly, whereas elevated blood pressure and/or increased plasma angiotensin II levels suppressed them. Human and mouse renal renin mRNA levels changed similarly but to a lesser degree. These studies demonstrate that human renin synthesis and secretion respond appropriately in 45-kb hREN mice to physiological stimuli. This most likely results from appropriate cell-specific expression of the transgene conferred by the extended transgene flanking sequences.
AIM: To study the effect of captopril (Cap) on platelet cytosolic free calcium concentration ([Ca2+]i), platelet aggregation (PAg), and plasma TXA2/PGI2 ratio in the renovascular hypertensive rats. METHODS: Blood pressure was measured once a week by tail-cuff microphonic manometer. Platelet [Ca2+]i was measured by Fura 2-AM. Plasma angiotensin II (Ang), thromboxane A2 (TXA2), and prostacycline (PGI2) were measured by radioimmunoassay. RESULTS: Platelet [Ca2+]i and PAg increased (P < 0.01), while plasma Ang and TXA2/PGI2 ratio elevated (P < 0.05) in the renovascular hypertensive rats; platelet [Ca2+]i and plasma TXA2/PGI2 ratio reduced markedly after i.g. Cap 100 mg.kg-1.d-1 compared with saline for 2 wk. CONCLUSION: The altered TXA2/PGI2 after Cap treatment contributed to the improvement of the platelet [Ca2+]i and PAg.
Immunoprotection against the infection by Leptospira interrogans serogroup Icterohemorrhagiae serovar lai strain 017 was demonstrated in guinea pigs vaccinated with DNA recombinant plasmid rpDJt and expressed protein P68 derived from genomic library of Leptospira strain 017. Thirty days after active immunization, each group received intraperitoneally (1/2 dose) and subcutaneously (1/2 dose) inoculum of L. interrogans serovar lai stain 017; cultures were adjusted to 5 x 10(8) cells/ml. All guinea pigs were observed for 10 days after challenge. Survival (%) of P68 group was 100(7/7); P23 group was 75(3/4); group rpDJt was 77(10/13); group lack recombinant (control) pT7-7 was 25 (3/12), and group with whole-cell inactivated vaccine was 93(13/14). Although the protective antigen in the leptospires has yet to be determined, it is evident that expressed protein P68 conferred a high degree of immunoprotection in guinea pigs.
OBJECTIVE: To observe the effects of Salvia Miltiorrhizae Composita (SMC) on blood gas variations of nitroglycerin (NTG) controlled hypotension. METHODS: Sixteen patients who were arranged to undergo operation under general anesthesia in controlled hypotension condition were divided randomly into control group (n = 8) and SMC group (treated group, n = 8), NTG were used to create controlled hypotension in both groups and blood pressure decrease about 30% comparing with the control values. Patients in the treated group were administered intravenously with SMC (16 ml) before the utilization of NTG. RESULTS: Significant PaO2 decrease and Pa-etCO2 increase were observed in the control group during the controlled hypotension period. However, blood gas values in the treated group did not evidently change. CONCLUSION: It suggested that SMC can prevent the side effects of NTG on blood gas.
We have isolated the human PEG1/MEST gene and have investigated its imprinting status and parental-specific methylation. FISH mapping assigned the gene to chromosome 7q32, and homologous sequences were identified on the short arm of human chromosomes 3 and 5. Through the use of a newly identified intragenic polymorphism, expression analysis revealed that PEG1/MEST is monoallelically transcribed in all fetal tissues examined. In two informative cases, expression was shown to be confined to the paternally derived allele. In contrast to the monoallelic expression observed in fetal tissues, biallelic expression was evident in adult blood lymphocytes. Biallelic expression in blood is supported by the demonstration of PEG1/MEST transcripts in a lymphoblastoid cell line with maternal uniparental disomy 7. The human PEG1/MEST gene spans a genomic region of approximately 13 kb. Sequence analysis of the 5' region of PEG1/MEST revealed the existence of a 620-bp-long CpG island that extends from the putative promoter region into intron 1. We demonstrate that this CpG island is methylated in a parent-of-origin-specific manner. All MspI/HpaII sites were unmethylated on the active paternal allele but methylated on the inactive maternal one.
Monobromobimane (mBBr) can label both Cys111 and Cys17 of rat liver glutathione S-transferase, 1-1 (GST 1-1). However, selective modification of Cys111 was achieved by the maleimide-based sulfhydryl reagents N-ethylmaleimide (NEM) and fluorescein 5-maleimide (NFM). Incubation of GST 1-1 with 5 mM NEM for 30 min at pH 7.5 and 25 degrees C leads to the formation of modified enzyme with 92% residual activity toward 1-chloro-2,4-dinitrobenzene and completely blocks Cys111 from subsequent reaction with either NFM or mBBr. Reaction of GST 1-1 with 0.2 mM NFM under the same conditions affords a modified enzyme with only 14% residual activity even though NFM and NEM target the same Cys111. The results indicate that when the bulky fluorescein is covalently bound to Cys111, the ligand projects into both the xenobiotic binding site and the glutathione site. After NEM or NFM modification of GST 1-1, the enzyme was further modified by monobromobimane at Cys17 with loss of activity. Together with the only tryptophan (Trp20), fluorescein linked to Cys111 and bimane to Cys17 provide three fluorescent probes to study the solution structure of GST 1-1. Fluorescence spectral analysis suggests that Trp20 and bimane linked to Cys17 are located in a relatively hydrophobic environment, while fluorescein linked to Cys111 is located in a charged environment. These fluorescent probes constitute three sets of donor-acceptor pairs for the measurement of fluorescence energy transfer, and distances calculated from such measurements are 20 A between Trp20 and bimane at Cys17, 19 A between Trp20 and fluorescein at Cys111, and < 22 A between bimane at Cys17 and fluorescein at Cys111. Molecular modeling studies indicate that fluorescein lies between the two subunits, is surrounded by charged residues, and is extended into the xenobiotic binding site. They also suggest that mBBr must approach from the dimer interface in order to reach the reaction site at Cys17.
The dynamics for the hydraulic process of calcium phosphate cement (CPC) were investigated by X-ray diffraction quantitative analysis. The results show that the hardening process of CPC is initially controlled by the dissolution of reactants in a 4-h period and subsequently by diffusion through the product layer of hydroxyapatite (HAP) around the grains. The compressive strength rises approximately linearly with the increase of the extent of conversion in a 4-h period, and a maximum compressive strength of about 51 MPa, which is superior to those reported by the references, is obtained in 4 h. Then the compressive strength drops a little with an increase in the extent of conversion. The final product of setting reaction is acicular HAP crystal. Crystal seed not only reduces the setting time but also drops the compressive strength. The variation of pH in CPC slurry from 7.5 to 10.5 reveals that the control step of the dissolution process in the hardening process is the dissolution of dicalcium phosphate anhydrous and the presence of crystal seed will reduce the supersaturation to produce HAP.
Monobromobimane (mBBr) is a substrate of both mu- and alpha-class rat liver glutathione S-transferases, with Km values of 0.63 microM and 4.9 microM for the mu-class isozymes 3-3 and 4-4, respectively, and 26 microM for the alpha-class isozymes 1-1 and 2-2. In the absence of substrate glutathione, mBBr acts as an affinity label of the 1-1 as well as mu-class isozymes, but not of the alpha-class 2-2 isozyme. Incubation of rat liver isozyme 1-1 with mBBr at pH 7.5 and 25 degrees C results in a time-dependent inactivation of the enzyme but at a slower (threefold) rate than for reactions with the mu-class isozyme 3-3 and 4-4. The rate of inactivation of 1-1 isozyme by mBBr is not decreased but, rather, is slightly enhanced by S-methyl glutathione. In contrast, 17 beta-estradiol-3,17-disulfate (500 microM) gives a 12.5-fold decrease in the observed rate constant of inactivation by 4 mM mBBr. When incubated for 60 min with 4 mM mBBr, the 1-1 isozyme loses 60% of its activity and incorporates 1.7 mol reagent/mol subunit. Peptide analysis after thermolysin digestion indicates that mBBr modification is equally distributed between two cysteine residues at positions 17 and 111. Modification at these two sites is reduced equally in the presence of the added protectant, 17 beta-estradiol-3,17-disulfate, suggesting that Cys 17 and Cys 111 reside within or near the enzyme's steroid binding sites. In contrast to the 1-1 isozyme, the other alpha-class isozyme (2-2) is not inactivated by mBBr at concentrations as high as 15 mM. The different reaction kinetics and modification sites by mBBr suggest that distinct binding site structures are responsible for the characteristic substrate specificities of glutathione S-transferase isozymes.
Although it is well known that exogenous nerve growth factor (NGF) can dramatically affect the phenotype of the basal forebrain cholinergic neurons in normal, aged, or lesioned animals, whether its actions are restricted to the terminal field level of these cholinergic neurons has yet to be established. In most cases, NGF has been applied into the cerebroventricle space giving it access to both the terminal fields and somatodendritic regions. The recent demonstration that TrkA, the essential component of high-affinity NGF receptors, is expressed not only at the distal fields (terminals and distal axons) but also at the proximal fields (cell bodies, dendrites, and proximal axons) of the basal forebrain cholinergic neurons has provoked renewed interest in this problem. More recently, it was further demonstrated that in Alzheimer's disease (AD), the NGF peptide increased throughout the brain but decreased in the nucleus basalis magnocellularis (NBM), suggesting that there is an impaired retrograde transport of NGF from the cortex to the NBM. Thus, it will be crucial to clarify whether or not the TrkA receptors on the somatodendritic fields of the NBM cholinergic neurons respond to exogenous NGF in order to support the rationale for site-directed neurotrophic factor therapy in AD or other neurological disorders. To clarify this issue, we delivered 2.5S purified mouse NGF locally into the cortex or corpus striatum adjacent to the NBM of naive and cortically devascularized mature male Wistar rats. The local distribution of exogenous NGF was demonstrated by immunohistochemical staining. In naive rats, an NGF dose of 84.00 or 16.80 microg infused into either the cortex or the corpus striatum for 2 weeks caused ipsilateral hypertrophy of the cholinergic neurons of the NBM. In cortically devascularized animals, an NGF dose of 84.00 microg delivered into the cortex and 84.00 microg or 16.80 microg infused into the striatum adjacent to the NBM for 2 weeks rescued the ipsilateral cholinergic phenotype of NBM neurons of the basolocortical pathway from retrograde degeneration. Thus, exogenous NGF can affect the cholinergic phenotype of the NBM regardless of whether it is presented to their nerve terminal fields or their somatodendritic region. The present results provide new evidence that the TrkA receptors present in the somatodendritic region of the cholinergic neurons of the NBM are functional and capable of modulating neuronal phenotype in the naive and lesioned CNS, when applied pharmacologically.
We previously characterized the rabbit recombination activating gene-2 (RAG-2) coding region and a portion of the cDNA. Rabbit RAG-2 mRNA, however, was shown to be approximately twice as large as the predominant form expressed in other vertebrate species, suggesting that it contained additional coding and/or untranslated regions (UTR). In this report, we map and sequence the complete 5' and 3' UTRs of the rabbit RAG-2 transcript and identify and sequence the genomic regions from which they are transcribed. The data show that, with the exception of a 300 nucleotide 5' UTR, almost all of the additional sequence belongs to the 3' UTR and that the 3' UTR sequence is transcribed from a single large exon that encodes most of the coding region and all of the 3' UTR. The 3' UTR contains four poly A signal sites, the last of which is closely followed by a GU-rich region. The rabbit 3' UTR has a high level of identity with the homologous region downstream of the human RAG-2 gene but not with the mouse RAG-2 gene. The region of identity extends several hundred nucleotides beyond the transcribed region and terminates in a series of dinucleotide (TG) repeats. The data are discussed in terms of RAG gene and 3' UTR function, regulation, and evolution.
In this study, the biocompatibility of calcium phosphate cement (CPC) was systematically investigated, which included systemic injection acute toxicity assay, cell culture cytotoxicity assay, gene mutation assay (Ames test), chromosome aberration assay (micronucleus test), DNA damage assay (unscheduled DNA synthesis test) and implant histological evaluation. The results showed that the CPC had no toxicity and all tests for mutagenicity and potential carcinogenicity of CPC extracts are negative. CPC was implanted into the femurs of rabbits under no-load condition and the histology of specimens revealed that the implant tightly joined with the surrounding bone, only a very slight histological inflammatory reaction occurred. Thus, CPC is a highly biocompatible material and seems to be safe for application in humans.
The neuronal organization and patterns of afferent innervation are abnormal in the cingulate cortex in schizophrenia, and associated changes in synaptic terminals could be present. A panel of monoclonal antibodies was defined with biochemical and fusion protein studies as detecting syntaxin (antibody SP6), synaptophysin (antibody SP4) and synaptosomal-associated protein-25 (antibody SP12). These antibodies and a polyclonal antibody reactive with neural cell adhesion molecule were used to investigate the cingulate cortex in schizophrenia. Immunocytochemistry indicated that syntaxin immunoreactivity had a considerably wider distribution than synaptophysin. Overall, multivariate analysis indicated increased synaptic terminal protein immunoreactivity in schizophrenia compared to controls (P=0.004). Controlled for age and post mortem interval, syntaxin immunoreactivity was significantly elevated in schizophrenia (P=0.004), and neural cell adhesion molecule immunoreactivity was also elevated (P=0.05). The neural cell adhesion molecule to synaptophysin ratio was increased (P=0.005), possibly indicating the presence of less mature synapses in schizophrenia. Elevated syntaxin immunoreactivity is consistent with increased glutamatergic afferents to the cingulate cortex in schizophrenia, and combined with the neural cell adhesion molecule to synaptophysin ratio results suggests that synaptic function in this region in schizophrenia may be abnormal.
mel-18 is a mammalian homolog of Drosophila melanogaster Polycomb group genes. Mice lacking the mel-18 gene show a posterior transformation of the axial skeleton, severe combined immunodeficiency, and a food-passing disturbance in the lower intestine due to hypertrophy of the smooth muscle layer. In this study, the severe combined immunodeficiency observed in mel-18 mutant mice is correlated with the impaired mitotic response of lymphocyte precursors upon interleukin-7 stimulation. Strikingly, the axial skeleton and lymphoid phenotypes are identical in both mel-18 and bmi-1 mutants, indicating that the Mel-18 and Bmi-1 gene products might act in the same genetic cascade. These results suggest that mammalian Polycomb group gene products are involved in cell cycle progression in the immune system.