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Biomedical subjects

J Huang

Publications and source records attributed to J Huang.

At least 91 records · Page 5Linked to original sources
PubMed2001

The allatostatin gene of the cricket Gryllus bimaculatus (Ensifera, Gryllidae).

The gene encoding allatostatins (AST) of the FGLamide family from the cricket Gryllus bimaculatus is expressed in the brain. The mRNA, which contains four polyadenylation signals, encodes a hormone precursor that is split into at least 14 putative hormones. Five of them have been previously found in the cricket, six to seven others, or their close homologues, are known from other insects. Hormone AST 2 contains an internal cleavage site and may exist in a shorter version 2b. The hormones AST 3 and 4 are identical. The cDNA sequence revealed that a single point mutation and a single deletion eliminated an additional hormone between AST 12 and 13. The deduced hormone precursor is very similar to that in cockroaches, but is different from a shorter precursor in locusts, indicating that the gene evolved very fast in the latter. Regions conserved between cockroaches and crickets include parts of the acidic spacers that separate clusters of hormones, suggesting that these spacers may have additional functions.

Animals↗
PubMed2001

Neural circuitry of the kidney: NO-containing neurons.

The neurons synthesizing nitric oxide (NO) that are part of the renal sympathetic pathways were located by double-staining for the neuronal isoform of nitric oxide synthase (nNOS) using immunocytochemistry to identify NO-synthesizing neurons and transneuronal tracing following infection of the left kidney with pseudorabies virus (PRV). Following kidney injection with PRV, the animals survived 4-day post-inoculation prior to sacrifice and tissue processing. PRV-infected neurons that double-stained for nNOS were found in the paraventricular hypothalamic nucleus (PVN), the raphe obscurus nucleus (ROb), the ventromedial medulla (VMM), the rostral ventrolateral medulla (rVLM) and the A5 cell group. In the thoracolumbar spinal cord, nNOS neurons co-localized with PRV-infected cells in the dorsal horn in laminae I, III-V ipsilateral to the injected kidney and in lamina X, the intermediolateral cell column, the lateral funiculus, the intercalated nucleus and the central autonomic area. We conclude that NO synthesizing cells may significantly affect renal autonomic pathways in the rat by interacting with the renal sensory and sympathomotor circuitry at multiple sites.

Afferent Pathways↗
PubMed2001

Amination reactions of aryl halides with nitrogen-containing reagents mediated by palladium/imidazolium salt systems.

Nucleophilic N-heterocyclic carbenes have been conveniently used as catalyst modifiers in amination reactions involving aryl chlorides, aryl bromides, and aryl iodides with various nitrogen-containing substrates. The scope of a coupling process using a Pd(0) or Pd(II) source and an imidazolium salt in the presence of a base, KO(t)Bu or NaOH, was tested using various substrates. The Pd(2)(dba)(3)/IPr.HCl (1, IPr = 1,3-bis(2,6-diisopropylphenyl)imidazol-2-ylidene) system presents the highest activity with respect to electron-neutral and electron-rich aryl chlorides. The ligand is also effective for the synthesis of benzophenone imines, which can be easily converted to the corresponding primary amines by acid hydrolysis. Less reactive indoles were converted to N-aryl-substituted indoles using as supporting ligand the more donating SIPr.HCl (5, SIPr = 1,3-bis(2,6-diisopropylphenyl)-4,5-dihydroimidazol-2-ylidene). The Pd(OAc)(2)/SIPr.HCl/NaOH system is efficient for the N-arylation of diverse indoles with aryl bromides. The general protocol developed has been applied successfully to the synthesis of a key intermediate in the synthesis of an important new antibiotic. Mechanistically, palladium-to-ligand ratio studies strongly support an active species bearing one nucleophilic carbene ligand.

Amination↗
PubMed2001

Chromatography on DEAE ion-exchange and Protein G affinity columns in tandem for the separation and purification of proteins.

A high-performance liquid-chromatographic method based on coupled DEAE anion-exchange and Protein G affinity columns has been developed for the simultaneous separation and purification of immunoglobulin G and albumin from mouse serum. The diluted mouse serum was injected directly into this system, and the proteins were eluted separately from the DEAE and Protein G columns, coupled in series, by the column-switching technique. The advantages of this method are that IgG and albumin can be separated and purified simultaneously, the expensive affinity column is protected from contamination by the impurities in the mouse serum, and it is fast, selective, robust, and reproducible.

Animals↗
PubMed2001

Insulin can regulate GLUT4 internalization by signaling to Rab5 and the motor protein dynein.

Insulin stimulates glucose transport by promoting translocation of the insulin-sensitive glucose transporter isoform 4 (GLUT4) from an intracellular compartment to the cell surface. This movement is accomplished by stimulation of GLUT4 exocytosis as well as inhibition of endocytosis. However, the molecular mechanisms for these effects remain unclear. In this study, we found that the GTP-binding protein Rab5 physically associated with the motor protein dynein in immunoprecipitants from both untransfected cells and cells transfected with GFP-Rab5 constructs. Microinjection of anti-Rab5 or anti-dynein antibody into 3T3-L1 adipocytes increased the basal level of surface GLUT4, did not change the insulin-stimulated surface GLUT4 level, and inhibited GLUT4 internalization after the removal of insulin. Photoaffinity labeling of Rab5 with [gamma-(32)P]GTP-azidoanilide showed that insulin inhibited Rab5-GTP loading. By using microtubule-capture assays, we found that insulin also caused a significant decrease in the binding of dynein to microtubules. Furthermore, pretreatment of cells with the PI3-kinase inhibitor LY294002 inhibited the effects of insulin on both Rab5-GTP loading and dynein binding to microtubules. In conclusion, these data indicate that insulin signaling inhibits Rab5 activity and the interaction of dynein with microtubules in a PI3-kinase-dependent manner, and that these effects may inhibit the rate of GLUT4 internalization. As such, our results present a previously uncharacterized insulin-signaling pathway involving Rab5, the motor protein dynein, and the cytoskeleton to regulate directional GLUT4 movement, facilitating GLUT4 distribution to the cell surface.

3T3 Cells↗
PubMed2001

Adoptive transfer of autologous Epstein-Barr virus-specific cytotoxic T cells for nasopharyngeal carcinoma.

Tumor cells from NPC patients are regularly and latently infected with EBV. To examine whether the virus serves as target for immune intervention of the cancer, we determined levels of EBV-specific CTLp in peripheral blood from NPC patients, long-term survivors of the cancer and healthy subjects. CTLp levels of test subjects varied between 3- 3,000/10(6) PBMCs. The plasma EBV burden increased when the CTLp level fell below 150, whereas the EBV burden of PBMCs was not correlated with CTLp level. Compared with healthy carriers, CTLp levels of patients were lower and varied over a wider range, between 3-1,500/10(6) PBMCs. The quantitative immune deficit was probably attributed to the tumor because, first, CTLp in survivors was restored to levels similar to those in healthy carriers after the tumor had been successfully treated. Second, the CTLp level changed as disease progressed, being lower in local disease, increased in locoregional disease and decreased again when the tumor metastasized. Based on these findings, 4 patients with advanced disease were infused with 5 x 10(7)-3 x 10(8) autologous EBV CTLs. The treatment was safe and unaccompanied by inflammatory or other complications, but whether it improved tumor control could not be discerned from the large tumor bulk. Nevertheless, the treatment regularly increased CTLp levels of patients, maintained it at higher levels for protracted periods and, in 3 patients, restored host surveillance of EBV replication, reducing the plasma EBV burden. Taken together, these results provided a rationale to further explore EBV as a target of immune intervention of NPC.

Adoptive Transfer↗
PubMed2001

Polyglutamine-expanded ataxin-7 antagonizes CRX function and induces cone-rod dystrophy in a mouse model of SCA7.

Spinocerebellar ataxia type 7 (SCA7) is an autosomal dominant disorder caused by a CAG repeat expansion. To determine the mechanism of neurotoxicity, we produced transgenic mice and observed a cone-rod dystrophy. Nuclear inclusions were present, suggesting that the disease pathway involves the nucleus. When yeast two-hybrid assays indicated that cone-rod homeobox protein (CRX) interacts with ataxin-7, we performed further studies to assess this interaction. We found that ataxin-7 and CRX colocalize and coimmunoprecipitate. We observed that polyglutamine-expanded ataxin-7 can dramatically suppress CRX transactivation. In SCA7 transgenic mice, electrophoretic mobility shift assays indicated reduced CRX binding activity, while RT-PCR analysis detected reductions in CRX-regulated genes. Our results suggest that CRX transcription interference accounts for the retinal degeneration in SCA7 and thus may provide an explanation for how cell-type specificity is achieved in this polyglutamine repeat disease.

Age Factors↗
PubMed2001

Role of the PHTH module in protein substrate recognition by Bruton's agammaglobulinemia tyrosine kinase.

Defects in Bruton's tyrosine kinase (Btk) are responsible for X chromosome-linked agammaglobulinemia in patients. Mutations in each of the structural domains of Btk have been detected in patients, yet a mechanistic explanation for most of these mutant phenotypes is lacking. To understand the possible role of the unique pleckstrin homology and Tec homology (PHTH) module of Btk, we have compared the enzymatic properties of full-length Btk and a Btk mutant lacking the PHTH module (BtkDeltaPHTH). Here we show that Btk and BtkDeltaPHTH have similar basal catalytic activity but very different abilities to recognize protein substrates. Furthermore, the catalytic domain of Btk is inactive, in contrast to the catalytic domain of the prototypical Src tyrosine kinase that retains full catalytic ability. These data suggest that the PHTH module plays an important role in protein substrate recognition, that Btk and Src likely have different interdomain organizations and regulations, and that alterations in substrate recognition might play a role in X chromosome-linked agammaglobulinemia.

Agammaglobulinaemia Tyrosine Kinase↗
PubMed2001

Dehydroascorbic acid, a blood-brain barrier transportable form of vitamin C, mediates potent cerebroprotection in experimental stroke.

Neuronal injury in ischemic stroke is partly mediated by cytotoxic reactive oxygen species. Although the antioxidant ascorbic acid (AA) or vitamin C does not penetrate the blood-brain barrier (BBB), its oxidized form, dehydroascorbic acid (DHA), enters the brain by means of facilitative transport. We hypothesized that i.v. DHA would improve outcome after stroke because of its ability to cross the BBB and augment brain antioxidant levels. Reversible or permanent focal cerebral ischemia was created by intraluminal middle cerebral artery occlusion in mice treated with vehicle, AA, or DHA (40, 250, or 500 mg/kg), either before or after ischemia. Given before ischemia, DHA caused dose-dependent increases in postreperfusion cerebral blood flow, with reductions in neurological deficit and mortality. In reperfused cerebral ischemia, mean infarct volume was reduced from 53% and 59% in vehicle- and AA-treated animals, respectively, to 15% in 250 mg/kg DHA-treated animals (P < 0.05). Similar significant reductions occurred in nonreperfused cerebral ischemia. Delayed postischemic DHA administration after 15 min or 3 h also mediated improved outcomes. DHA (250 mg/kg or 500 mg/kg) administered at 3 h postischemia reduced infarct volume by 6- to 9-fold, to only 5% with the highest DHA dose (P < 0.05). In contrast, AA had no effect on infarct volumes, mortality, or neurological deficits. No differences in the incidence of intracerebral hemorrhage occurred. Unlike exogenous AA, DHA confers in vivo, dose-dependent neuroprotection in reperfused and nonreperfused cerebral ischemia at clinically relevant times. As a naturally occurring interconvertible form of AA with BBB permeability, DHA represents a promising pharmacological therapy for stroke based on its effects in this model of cerebral ischemia.

Animals↗
PubMed2001

Risk of thyroid carcinoma in a female population after radiotherapy for breast carcinoma.

BACKGROUND: There is increasing concern regarding the risk of developing a second primary tumor in adjacent organs as a result of scattered radiation among patients who have undergone radiotherapy (RT) for breast carcinoma. Previous studies have focused mainly on the possible increase in the incidence of contralateral breast carcinoma. To the authors' knowledge, the risk of thyroid carcinoma among these women has not been explored to date. METHODS: In this population-based, retrospective cohort study, the authors identified 194,798 women who were diagnosed with invasive breast carcinoma (exclusive of those with distant metastasis) between 1973 and 1993, and ascertained subsequent cases of thyroid carcinoma utilizing data from the Surveillance, Epidemiology, and End Results (SEER) program of the U.S. National Cancer Institute. Poisson regression was used to calculate the age-standardized incidence ratio (SIR) of thyroid carcinoma and to model the influence of RT on the relative risk (RR) between the RT cohort (48,495 women) and the non-RT cohort (146,303 women). RESULTS: A total of 28 women in the RT cohort and 112 women in the non-RT cohort subsequently developed thyroid carcinoma. The distribution of thyroid carcinoma histologies in both the RT cohort and the non-RT cohort was similar to that in the female general population. Overall, there was no significant increase in the risk of thyroid carcinoma in either the RT cohort or the non-RT cohort compared with the general population; the SIR was 1.1 (95% confidence interval [95% CI], 0.8-1.6) for the RT cohort and 1.2 (95% CI, 1.0-1.4) for the non-RT cohort. When the RT cohort was compared with the non-RT cohort, the RR of thyroid carcinoma was 1.0 (95%CI, 0.7-1.5). CONCLUSIONS: The risk of radiation-associated thyroid carcinoma after initial RT for breast carcinoma was so low as to be undetectable in the current large population-based study. Continued monitoring of these women will be required to document that these findings are maintained with even longer follow-up periods. However, with 10,895 women having been followed for > 10 years at the time of last follow-up in the current study, these findings should be reassuring to women considering RT for their breast carcinoma. Therefore, women who have received RT for breast carcinoma require no special surveillance for their thyroid gland. Furthermore, previous breast radiation need not be a factor in determining the optimal management of thyroid nodules arising in women who received RT for breast carcinoma.

Adenocarcinoma, Follicular↗
PubMed2001

beta -Arrestin-mediated recruitment of the Src family kinase Yes mediates endothelin-1-stimulated glucose transport.

The insulin and the endothelin type A (ETA) receptor both can couple into the heterotrimeric G protein alpha(q/11) (Galpha(q/11)), leading to Galpha(q/11) tyrosine phosphorylation, phosphatidylinositol 3-kinase activation, and subsequent stimulation of glucose transport. In this study, we assessed the potential role of Src kinase in ET-1 signaling to glucose transport in 3T3-L1 adipocytes. Src kinase inhibitor PP2 blocked ET-1-induced Src kinase activity, Galpha(q/11) tyrosine phosphorylation, and glucose transport stimulation. To determine which Src family kinase member was involved, we microinjected anti-c-Src, -c-Fyn, or -c-Yes antibody into these cells and found that only anti-c-Yes antibody blocked GLUT4 translocation (70% decreased). Overexpression or microinjection of a dominant negative mutant (K298M) of Src kinase also inhibited ET-1-induced Galpha(q/11) tyrosine phosphorylation and GLUT4 translocation. In co-immunoprecipitation experiments, we found that beta-arrestin 1 associated with the ETA receptor in an agonist-dependent manner and that beta-arrestin 1 recruited Src kinase to a molecular complex that included the ETA receptor. Microinjection of beta-arrestin 1 antibody inhibited ET-1- but not insulin-stimulated GLUT4 translocation. In conclusion, 1) the Src kinase Yes can induce tyrosine phosphorylation of Galpha(q/11) in response to ET-1 stimulation, and 2) beta-arrestin 1 and Src kinase form a molecular complex with the ETA receptor to mediate ET-1 signaling to Galpha(q/11) with subsequent glucose transport stimulation.

3T3 Cells↗
PubMed2001

Effects of selective cyclooxygenase-2 inhibition on vascular responses and thrombosis in canine coronary arteries.

BACKGROUND: Prostanoid synthesis via the action of cyclooxygenase-2 (COX-2) is a component of the inflammatory response. Prostacyclin, a product of COX-2 in vascular endothelium, has important physiological roles, such as increasing blood flow to injured tissues, reducing leukocyte adherence, and inhibiting platelet aggregation. We examined the possibility that selective COX-2 inhibition could suppress the protective effects of prostacyclin, resulting in an alteration of the hemostatic balance and vascular tone. METHODS AND RESULTS: Circumflex coronary artery thrombosis was induced in dogs by vascular electrolytic injury. Orally administered celecoxib (COX-2 inhibition) or high-dose aspirin (HDA) (COX-1 and COX-2 inhibition) did not alter time to occlusive thrombus formation compared with controls (celecoxib 77.7+/-7.2 minutes, HDA 72.0+/-18.5 minutes, control 93.0+/-21.8 minutes). Oral HDA with an endothelial recovery period (HDA-ER) (COX-1 inhibition) produced a significant increase in time to vessel occlusion (257.0+/-41.6 minutes). The observed increase in time to occlusion was abolished when celecoxib was administered to animals dosed with HDA-ER (80.7+/-20.6 minutes). The vasomotor effect of endothelium-derived prostacyclin was examined by monitoring coronary flow during intracoronary administration of arachidonic acid or acetylcholine. In celecoxib-treated animals, vasodilation in response to arachidonic acid was reduced significantly compared with controls. CONCLUSIONS: The results indicate important physiological roles for COX-2-derived prostacyclin and raise concerns regarding an increased risk of acute vascular events in patients receiving COX-2 inhibitors. The risk may be increased in individuals with underlying inflammatory disorders, including coronary artery disease.

Acetylcholine↗
PubMed2001

Multiple cleavage activities of endonuclease V from Thermotoga maritima: recognition and strand nicking mechanism.

Endonuclease V is a deoxyinosine 3'-endonuclease which initiates removal of inosine from damaged DNA. A thermostable endonuclease V from the hyperthermophilic bacterium Thermotoga maritima has been cloned and expressed in Escherichia coli. The DNA recognition and reaction mechanisms were probed with both double-stranded and single-stranded oligonucleotide substrates which contained inosine, abasic site (AP site), uracil, or mismatches. Gel mobility shift and kinetic analyses indicate that the enzyme remains bound to the cleaved inosine product. This slow product release may be required in vivo to ensure an orderly process of repairing deaminated DNA. When the enzyme is in excess, the primary nicked products experience a second nicking event on the complementary strand, leading to a double-stranded break. Cleavage at AP sites suggests that the enzyme may use a combination of base contacts and local distortion for recognition. The weak binding to uracil sites may preclude the enzyme from playing a significant role in repair of such sites, which may be occupied by uracil-specific DNA glycosylases. Analysis of cleavage patterns of all 12 natural mismatched base pairs suggests that purine bases are preferrentially cleaved, showing a general hierarchy of A = G > T > C. A model accounting for the recognition and strand nicking mechanism of endonuclease V is presented.

Apurinic Acid↗
PubMed2001

Encapsulation in biodegradable microparticles enhances serum antibody response to parenterally-delivered beta-amyloid in mice.

Poly(lactide-co-glycolide) (PLG) microspheres were tested as a parenteral delivery system for human beta-amyloid (1-42) (Abeta), a potential immunotherapeutic undergoing assessment in Phase 1 studies for Alzheimer's disease (AD). Abeta was successfully encapsulated in PLG microspheres of average sizes of 3 or 15 microm diameter. Swiss Webster (SW) mice were injected by the sub-cutaneous (s.c.) or intra-peritoneal (i.p.) routes with 3-33 microg Abeta. Abeta-PLG microparticles (3 microm) induced dose-dependent antibody responses, which were maximal at 33 microg Abeta, while Abeta in phosphate-buffered saline (PBS) produced weak antibody responses at the same doses by both routes. Significantly increased antibody responses were seen for both small and large particle formulations given by the i.p. route in comparison to the s.c route. It was previously reported that passive immunisation with Abeta-specific antibodies cleared amyloid plaques in a mouse model of AD (Bard F, Cannon C, Barbour R, et al. Peripherally administered antibodies against amyloid beta-peptide enter the nervous system and reduce pathology in a mouse model of Alzheimer disease. Nature Med 2000;6:916-19), an indication that induction of serum antibody is a prerequisite for efficacy.

Amyloid beta-Peptides↗
PubMed2001

Evidence supporting WNT2 as an autism susceptibility gene.

We examined WNT2 as a candidate disease gene for autism for the following reasons. First, the WNT family of genes influences the development of numerous organs and systems, including the central nervous system. Second, WNT2 is located in the region of chromosome 7q31-33 linked to autism and is adjacent to a chromosomal breakpoint in an individual with autism. Third, a mouse knockout of Dvl1, a member of a gene family essential for the function of the WNT pathway, exhibits a behavioral phenotype characterized primarily by diminished social interaction. We screened the WNT2 coding sequence for mutations in a large number of autistic probands and found two families containing nonconservative coding sequence variants that segregated with autism in those families. We also identified linkage disequilibrium (LD) between a WNT2 3'UTR SNP and our sample of autism-affected sibling pair (ASP) families and trios. The LD arose almost exclusively from a subgroup of our ASP families defined by the presence of severe language abnormalities and was also found to be associated with the evidence for linkage to 7q from our previously published genomewide linkage screen. Furthermore, expression analysis demonstrated WNT2 expression in the human thalamus. Based on these findings, we hypothesize that rare mutations occur in the WNT2 gene that significantly increase susceptibility to autism even when present in single copies, while a more common WNT2 allele (or alleles) not yet identified may exist that contributes to the disorder to a lesser degree.

Amino Acid Sequence↗
PubMed2001

Increased risk of soft tissue sarcoma after radiotherapy in women with breast carcinoma.

BACKGROUND: Numerous studies to date have suggested an association between radiation exposure and the development of soft tissue sarcoma. The current study was performed to quantify the risk of soft tissue sarcoma in the vicinity of previously irradiated anatomic regions in women with breast carcinoma. METHODS: In this population-based, retrospective cohort study, 194,798 women who were diagnosed with invasive breast carcinoma (exclusive of those with distant metastasis) between 1973--1995 were identified, and subsequent soft tissue sarcoma cases utilizing the data from the Surveillance, Epidemiology, and End Results Program (SEER) were ascertained. Poisson regression analysis was used to calculate age standardized incidence ratios (SIR) and to model the influence of radiotherapy (RT) on the relative risk (RR) between the RT and non-RT cohorts. RESULTS: A total of 54 women in the RT cohort and 81 women in the non-RT cohort subsequently developed soft tissue sarcoma. In the RT cohort, the SIR was 26.2 (95% confidence interval [95% CI], 16.5--41.4) for angiosarcoma and was 2.5 (95% CI, 1.8--3.5) for other sarcomas; in the non-RT cohort, the SIRs were 2.1 (95% CI, 1.0--4.4) and 1.3 (95% CI, 1.0--1.7), respectively. The RT cohort demonstrated a higher risk of developing both angiosarcoma (RR: 15.9; 95% CI, 6.6--38.1) and other sarcomas (RR: 2.2; 95% CI, 1.4--3.3) compared with the non-RT cohort, and the largest increase was observed in the chest wall/breast. The elevated RR was significant even within 5 years of RT, but it reached a maximum between 5--10 years. CONCLUSIONS: The risk of soft tissue sarcoma, especially angiosarcoma, was elevated after RT in women with breast carcinoma.

Adult↗
PubMed2001

Molecular cloning and characterization of chemokine-like factor 1 (CKLF1), a novel human cytokine with unique structure and potential chemotactic activity.

Cytokines are small proteins that have an essential role in the immune and inflammatory responses. The repertoire of cytokines is becoming diverse and expanding. Here we report the identification and characterization of a novel cytokine designated as chemokine-like factor 1 (CKLF1). The full-length cDNA of CKLF1 is 530 bp long and a single open reading frame encoding 99 amino acid residues. CKLF1 bears no significant similarity to any other known cytokine in its amino acid sequence. Expression of CKLF1 can be partly inhibited by interleukin 10 in PHA-stimulated U937 cells. Recombinant CKLF1 is a potent chemoattractant for neutrophils, monocytes and lymphocytes; moreover, it can stimulate the proliferation of murine skeletal muscle cells. These results suggest that CKLF1 might have important roles in inflammation and in the regeneration of skeletal muscle.

Amino Acid Sequence↗