Search PubMed⌕ Search

Biomedical subjects

J Mao

Publications and source records attributed to J Mao.

At least 19 recordsLinked to original sources
PubMed2006

Evidence for a distinct group of nestin-immunoreactive neurons within the basal forebrain of adult rats.

Nestin is an intermediate filament protein serving as a marker for neuroprogenitor and stem cells. Here we report that a cluster of previously unrecognized nestin immunoreactive (nestin-ir) neurons was located in the medial septum-diagonal band of Broca (MS-DBB) of the basal forebrain in adult rats. Nestin-ir neurons were exclusively located in the MS-DBB and intermingled with choline acetyltransferase-ir (ChAT-ir), parvalbumin-ir (PV-ir), or nicotinamide adenine dinucleotide phosphate (NADPH)-diaphorase reactive (NADPHd-reactive) neurons. However, there was no colocalization between nestin-ir and PV-ir in single neurons in MS-DBB; only about 35% of nestin-ir neurons were ChAT-ir, and 8%-12% of nestin-ir neurons were NADPHd-reactive. Morphologically, nestin-ir neurons showed a larger size of somata than that of ChAT-ir or PV-ir neurons and the distribution of nestin-ir neurons spread across the rostro-caudal extent of the MS-DBB. Moreover, retrograde tracing revealed that a significant portion of these nestin-ir neurons projected to the thalamus and hippocampus. These results, for the first time, provide strong evidence that there exists a cluster of previously unrecognized nestin-ir neurons in MS-DBB of the basal forebrain in adult rats and that these nestin-ir neurons are distinguishable from ChAT-ir, PV-ir, and NADPHd-reactive neurons.

Acetylcholine↗
PubMed2005

Evidence for emergence of an amphibian iridoviral disease because of human-enhanced spread.

Our understanding of origins and spread of emerging infectious diseases has increased dramatically because of recent applications of phylogenetic theory. Iridoviruses are emerging pathogens that cause global amphibian epizootics, including tiger salamander (Ambystoma tigrinum) die-offs throughout western North America. To explain phylogeographical relationships and potential causes for emergence of western North American salamander iridovirus strains, we sequenced major capsid protein and DNA methyltransferase genes, as well as two noncoding regions from 18 geographically widespread isolates. Phylogenetic analyses of sequence data from the capsid protein gene showed shallow genetic divergence (< 1%) among salamander iridovirus strains and monophyly relative to available fish, reptile, and other amphibian iridovirus strains from the genus Ranavirus, suggesting a single introduction and radiation. Analysis of capsid protein sequences also provided support for a closer relationship of tiger salamander virus strains to those isolated from sport fish (e.g. rainbow trout) than other amphibian isolates. Despite monophyly based on capsid protein sequences, there was low genetic divergence among all strains (< 1.1%) based on a supergene analysis of the capsid protein and the two noncoding regions. These analyses also showed polyphyly of strains from Arizona and Colorado, suggesting recent spread. Nested clade analyses indicated both range expansion and long-distance colonization in clades containing virus strains isolated from bait salamanders and the Indiana University axolotl (Ambystoma mexicanum) colony. Human enhancement of viral movement is a mechanism consistent with these results. These findings suggest North American salamander ranaviruses cause emerging disease, as evidenced by apparent recent spread over a broad geographical area.

Animals↗
PubMed2004

Effect of epidermal growth factor and insulin-like growth factor I on porcine preantral follicular growth, antrum formation, and stimulation of granulosal cell proliferation and suppression of apoptosis in vitro.

The object of this study was to investigate the role of epidermal growth factor (EGF) and IGF-I in the regulation of preantral follicular growth, antrum formation, and granulosal cell proliferation/ apoptosis. Porcine preantral follicles were manually dissected and cultured for up to 8 d in Waymouth's (Exp. 1) or alpha-minimum Eagle's essential medium (Exp. 2 and 3) supplemented with 10 microg/mL of transferrin, 100 microg/mL of L-ascorbic acid, and 2 mU/mL of ovine FSH, in the presence (Exp. 1 and 3) or absence (Exp. 2) of 7.5% fetal calf serum. According to the experimental protocol, IGF-I (0, 1, 10, or 100 ng/mL; Exp. 1), or IGF-I (50 ng/mL), EGF (10 ng/mL) and EGF+IGF-I (Exp. 2 and 3) were added to the culture media. In Exp. 1, follicles exhibited a concentration-dependent response (P < 0.05) to IGF-I, with the highest rates of granulosal cell proliferation, follicular integrity, and recovery rate of cumulus cell-oocyte complexes and lowest incidence of apoptosis occurring at the highest IGF-I dose. In Exp. 2 serum-free medium, granulosal cell proliferation was low (1 to 5%), irrespective of whether EGF and/or IGF-I were present and cellular apoptosis was increased (P < 0.05) on d 4 and 8 in the EGF+IGF-I group compared with the addition of either factor alone. In Exp. 3, granulosal cell proliferation was high in all follicles cultured in serum-containing medium for the first 3 d, but fell sharply (P < 0.05) on d 4, except in media containing IGF-I. Collectively, EGF and IGF-I increased granulosal cell proliferation, decreased apoptosis, and promoted follicular antrum formation. These results may provide useful information for developing a preantral follicular culture system in which the oocytes are capable of fertilization and embryonic development.

Animals↗
PubMed2003

Oviduct-specific glycoprotein modulates sperm-zona binding and improves efficiency of porcine fertilization in vitro.

Oviduct-specific glycoprotein (OGP) displays estrus-associated regional and temporal differences in expression and localizes to the zona pellucida, perivitelline space, and plasma membrane of oviductal oocytes and embryos, suggesting that it may have a role in regulation of fertilization and/or early embryonic development. The aims of this study were to evaluate the effect of exogenous OGP on in vitro fertilization (IVF) and embryo development in the pig using a defined serum-free culture system. In vitro-matured porcine oocytes were incubated with homologous OGP (0, 1, 10, 20, and 40 microg/ml) for 3 h and then washed prior to IVF. Exposure of oocytes to 10 or 20 microg/ml porcine OGP (pOGP) significantly reduced the incidence of polyspermy compared with the control (P < 0.01) while maintaining high penetration rates. When oocytes, spermatozoa, or both were preincubated with 10 microg/ml pOGP prior to IVF, the incidence of polyspermy was similarly reduced (P < 0.01) by all three treatments without affecting penetration rates. The ability of spermatozoa to undergo calcium ionophore-induced acrosome reaction was similar with or without exposure to pOGP. However, significantly fewer spermatozoa (P < 0.01) bound to the zona pellucida when oocytes were preincubated with pOGP. To evaluate the effect of pOGP on embryo development, embryos were cultured in pOGP-supplemented medium for 48 h or 144 h. Both transient and continuous exposure to pOGP significantly enhanced cleavage and blastocyst formation rate compared with the control (P < 0.01). These data demonstrate that exposure of either in vitro-matured oocytes or spermatozoa to pOGP decreased polyspermy and spermatozoa binding while maintaining high penetration rates of pig oocytes fertilized in vitro. Furthermore, pOGP exerted an embryotrophic effect independent of effects demonstrated on spermatozoa and oocytes at fertilization.

Animals↗
PubMed2003

A threonine residue (Thr71) at the intracellular end of the M1 helix plays a critical role in the gating of Kir6.2 channels by intracellular ATP and protons.

ATP-sensitive K+ (K(ATP)) channels are known to be gated by several intracellular molecules, but the gating mechanisms remain unclear. To understand the relationship of channel gating to ligand binding, we studied Kir6.2 channel gating by ATP and protons, which inhibit and activate the channel, respectively. We have previously shown that a threonine residue (Thr71) is critical for the pH sensitivity of Kir6.2 channel. If this site is involved in channel gating rather than ligand binding, it should affect channel gating by both ATP and proton. To test this hypothesis we performed a mutation analysis. Site-specific mutations of Thr71 to a bulky residue reduced the ATP sensitivity by >100-fold and eliminated the pH sensitivity. Single-channel activity of these mutants was stabilized at the open state with no detectable rundown. Mutations to a small amino acid had little effect on the ATP and pH sensitivities. Mutations to intermediate amino acids reduced but did not abolish the ATP and pH sensitivities. Hydrophobicity is not critical, as both polar and nonpolar amino acids are found in each group. Mutation to a positively charged lysine markedly exacerbated the pH- but not ATP-sensitivity, whereas mutation to glutamate moderately reduced ATP and pH sensitivities. These results indicate that the residue mass is critical for Kir6.2 channel gating, a mass that should be below 120 daltons with no charge. The existence of such a site as Thr71 involved in channel gating by both ATP and proton suggests that channel gating in the K(ATP) channel likely is separate from ligand binding.

Adenosine Triphosphate↗
PubMed2003

Chromosomal abnormalities in Day-6, in vitro-produced pig embryos.

A cytogenetic study was undertaken to quantify, by chromosomal karyotyping, the incidence and type of chromosomal abnormalities present in Day-6 in vitro-produced (IVP) porcine embryos. Morphologically normal Day-6 blastocysts (n=318) were fixed and grouped into six classes according to the number of total cells (from < or =20 to 61-70). Of 248 embryos suitable for analysis, 97 (39.1%) displayed chromosomal abnormalities. The abnormalities included haploidy (9.3%), polyploidy (71.1%) and mixoploidy (19.6%). Within polyploid embryos, triploidy and tetraploidy showed the highest incidence (56.5 and 27.5%, respectively); among mixoploid embryos, diploid-triploid embryos (2n/3n) were prevalent (36.8%). Overall, the mean cell number was 34.3 +/- 12.1 and the mitotic index was 8.6 +/- 6.1. Chromosomally abnormal embryos had fewer (P<0.01) total cells compared to normal (2n) embryos (31.8 +/- 1.3 versus 35.9 +/- 1.0). In addition, the incidence of polyploidy decreased as the number of cells increased, while that of mixoploidy did not differ. These data indicate that polyploidy affects a large percentage of IVP porcine embryos capable of developing to blastocysts and the incidence of chromosomal abnormalities is much higher than that reported previously in in vivo embryos in this species. Given the ability of morphologically normal embryos with an abnormal chromosome complement to undergo preimplantation development in vitro, and the inability to identify blastocysts with abnormal karyotype without cytogenetic analysis, careful consideration should be given to factors affecting ploidy of IVP embryos, especially the incidence of polyspermic fertilization, when evaluating criteria of a porcine in vitro embryo production scheme.

Animals↗
PubMed2003

Ethical issues related to the use of placebo in clinical trials.

Controversy exists regarding the ethics of using placebo control groups in clinical trials when effective treatments exist. The debate was fuelled by the announcement of the fifth revision of the Declaration of Helsinki in 2000. This study reviews the history and scientific background surrounding the controversy and investigates the prevailing attitudes of Hong Kong researchers regarding this issue. The controversy has centred on several issues. The first involves the methodological superiority of placebo-controlled trials in discerning treatment effects. Secondly, it is unclear whether the treatment effects encompass absolute treatment effects (including placebo effects) or are confined to treatment-specific effects (excluding placebo effects). Thirdly, there are concerns that subjects in the placebo group could be exposed to a high risk for developing serious adverse events. Fourthly, it is unclear whether the standard of best available treatment should be local or international. Preliminary research findings suggest that the opinions of Hong Kong researchers are divided on the use of placebo control groups in clinical trials when effective treatment exists. Further research on the topic is therefore warranted, and training and consensus meeting may be necessary to minimise the confusion related to this issue.

Clinical Trials as Topic↗
PubMed2002

Protons activate homomeric Kir6.2 channels by selective suppression of the long and intermediate closures.

The ATP-sensitive K+ channels (KATP) play an important role in regulating membrane excitability. These channels are regulated by H+ in addition to ATP, ADP, and phospholipids. To understand how protons affect the single-channel properties, Kir6.2DeltaC36 currents were studied in excised inside-out patches. We chose to study the homomeric Kir6.2 channel with 36 amino acids deleted at the C-terminal end, as there are ADP/ATP-binding sites in the SUR subunit, which may obscure the understanding of the channel-gating process. In the absence of ATP, moderate intracellular acidosis (pH 6.8) augmented P(open) with small suppression (by approximately 10%) of the single-channel conductance. The long and intermediate closures were selectively inhibited, leading to a shortening of the mean closed time without significant changes in the mean open time. Stronger acidification (<pH6.2) caused channel rundown. Although similar changes in the single-channel properties were observed in the presence of 1 mM ATP, the P(open)-pH relationship curve was shifted by 0.17 pH units toward lower pH levels. ATP had no effect on the inhibition of single-channel conductance during acidosis. Mutation of His175 eliminated the pH effect on the single-channel kinetics, while the single-channel response to acidic pH was retained in the K185E mutant that greatly reduces the ATP sensitivity. These results indicate that Kir6.2DeltaC36 channel gating by protons and ATP relies on two distinct mechanisms opposite to each other, although the pH sensitivity is modulated by ATP.

Adenosine Triphosphate↗
PubMed2002

STAT5 binding contributes to lactational stimulation of promoter III expressing the bovine acetyl-CoA carboxylase alpha-encoding gene in the mammary gland.

Activity of acetyl-CoA carboxylase (ACC)-alpha is rate limiting for de novo synthesis of fatty acids. The encoding gene is expressed by three different promoters. We characterized promoter III (PIII) from cow, previously only known from sheep. Quantitation of transcripts by RNAse protection assays and real time PCR revealed that PIII is primarily expressed and strongly induced ( approximately 28-fold) in the lactating mammary gland. PIII transcripts are expressed in mammary epithelial cells (MEC) as shown by in situ hybridization. A 2999 bp segment of the PIII promoter conferred prolactin and dexamethasone inducibility to a luciferase reporter gene in stably transfected mouse MEC cells. Lactogenic induction was abolished if a unique signal transducer and activator of transcription (STAT)-binding site at position -797 was inactivated by two point mutations. An oligonucleotide probe harboring this STAT-site specifically bound nuclear proteins from the lactating mammary gland. Binding was abolished by those two point mutations and super-shift analyses showed that STAT5A factors are present in this complex. Hence, prolactin, acting through STAT5, contributes to the activation of ACC expression in the milk producing cells of the lactating mammary gland. We discuss that STAT5 might be important in determining the milk composition by coordinating fatty acid and protein synthesis during lactation.

Acetyl-CoA Carboxylase↗
PubMed2001

Characterization of an iridovirus from the cultured pig frog Rana grylio with lethal syndrome.

Three virus isolates, RGV-9506, RGV-9807 and RGV-9808, were obtained from cultured pig frogs Rana grylio undergoing lethal infections. Previously, the first isolate, RGV-9506, was shown to be an iridovirus based on ultrastructural and morphological studies. In the present study, the original isolate, along with 2 recent ones, were more extensively characterized by experimental infection studies, histopathology, electron microscopy, serological reactivity, gel electrophoresis of viral polypeptides and DNA restriction fragments, PCR amplification, and nucleic acid sequence analysis of the major capsid protein (MCP) gene. The 3 isolates were shown to be identical to each other, and very similar to FV3, the type species of the genus Ranavirus (family Iridoviridae). These results suggest that RGV should be considered a strain of FV3, and indicate that FV3-like iridoviruses are capable of causing widespread, severe disease among cultured frogs.

Amino Acid Sequence↗
PubMed2001

Biphasic effects of orchidectomy on calcitonin gene-related peptide synthesis and release.

We hypothesize that the decline of male gonadal hormones may play a role in age-related decrease of calcitonin gene-related peptide (CGRP) synthesis and release. Orchidectomized rats were raised with or without testosterone replacement and CGRP levels in serum and some tissues as well as the perfusate from the isolated mesenteric arterial bed (MAB) were measured at 1, 2 and 4 months after orchidectomy. CGRP levels of serum and tissues, and CGRP release from MAB were significantly elevated after 1 month and decreased after 4 months in orchidectomized rats. The changes were restored by testosterone replacement. Our results indicate that the age-related decline of testosterone might contribute to the age-related decrease of CGRP synthesis and release.

Animals↗
PubMed2001

Pharmaceutical development and manufacturing of a parenteral formulation of a novel antitumor agent, VNP40101M.

The objective of this study was to develop and manufacture a stable parenteral formulation for Phase I clinical trials of VNP40101M (1,2-Bis(methylsulfonyl)-1-(2-chloroethyl)-2-[(2-methylamino)carbonyl] hydrazine), a novel antitumor agent. The solubility and stability of the drug was determined. Solubility studies suggested that VNP40101M exhibited poor aqueous solubility but showed appreciable solubility in nonaqueous solvents. The aqueous solubility of the drug could not be increased by adjusting the pH. At a pH above 7, base-catalyzed decomposition of VNP40101M occurred. The low octanol-water partition coefficient of 0.75 suggested poor solubility in lipophilic solvents. Based on these preformulation observations, a parenteral formulation containing 10 mg/mL of VNP40101M was prepared in a solvent system consisting of 30% ethyl alcohol and 70% polyethylene glycol-300 (PEG-300). To minimize base-catalyzed hydrolytic degradation, citric acid at 0.6% concentration was included to acidify the formulation. Rubber closures, filter membranes, and liquid transfer tubing were selected on the basis of compatibility studies and absence of loss of drug due to adsorption of these components. The formulation was subjected to accelerated stability studies and dilution studies with large volume parenteral (LVP) solutions, normal saline, and 5% dextrose injection (D5W). The results of the dilution study indicated that the formulation could be diluted in these solutions up to 2 mg/mL for 8 hours without drug precipitation and degradation. Accelerated stability studies suggested that the product should be kept at 2 degrees C to 8 degrees C for long-term storage. The developed formulation was successfully scaled up and manufactured for use in clinical trials.

Adsorption↗
PubMed2001

Distinct histidine residues control the acid-induced activation and inhibition of the cloned K(ATP) channel.

The modulation of K(ATP) channels during acidosis has an impact on vascular tone, myocardial rhythmicity, insulin secretion, and neuronal excitability. Our previous studies have shown that the cloned Kir6.2 is activated with mild acidification but inhibited with high acidity. The activation relies on His-175, whereas the molecular basis for the inhibition remains unclear. To elucidate whether the His-175 is indeed the protonation site and what other structures are responsible for the pH-induced inhibition, we performed these studies. Our data showed that the His-175 is the only proton sensor whose protonation is required for the channel activation by acidic pH. In contrast, the channel inhibition at extremely low pH depended on several other histidine residues including His-186, His-193, and His-216. Thus, proton has both stimulatory and inhibitory effects on the Kir6.2 channels, which attribute to two sets of histidine residues in the C terminus.

Amino Acid Sequence↗
PubMed2001

Genomic distribution of three promoters of the bovine gene encoding acetyl-CoA carboxylase alpha and evidence that the nutritionally regulated promoter I contains a repressive element different from that in rat.

The enzyme acetyl-CoA carboxylase alpha (ACC-alpha) is rate-limiting for the synthesis of long-chain fatty acids de novo. As a first characterization of the bovine gene encoding this enzyme, we established the entire bovine ACC-alpha cDNA sequence (7041 bp) and used experiments with 5' rapid amplification of cDNA ends to determine the heterogeneous composition of 5' untranslated regions, as expressed from three different promoters (PI, PII and PIII). The individual locations of these promoters have been defined within an area comprising 35 kbp on Bos taurus chromosome 19 ('BTA19'), together with the segmentation of the first 14 exons. Primer extension analyses reveal that the nutritionally regulated PI initiates transcription from at least four sites. PI transcripts are much more abundant in adipose and mammary-gland tissues than in liver or lung. A 2.6 kb promoter fragment drives the expression of reporter genes only weakly in different model cells, irrespective of stimulation with insulin or dexamethasone. Thus bovine PI is basically repressed, like its analogue from rat. Finely graded deletions of PI map two separate elements, which have to be present together in cis to repress bovine PI. The distal component resides within a well-preserved Art2 retroposon element. Thus sequence, structure and evolutionary origin of the main repressor of PI in bovines are entirely different from its functional counterpart in rat, which had been identified as a (CA)(28) microsatellite. We show that, in different mammalian species, unrelated genome segments of different origins have been recruited to express as functionally homologous PI the ancient and otherwise highly conserved ACC-alpha-encoding gene.

Acetyl-CoA Carboxylase↗
PubMed2001

Requirement of multiple protein domains and residues for gating K(ATP) channels by intracellular pH.

ATP-sensitive K(+) channels (K(ATP)) are regulated by pH in addition to ATP, ADP, and phospholipids. In the study we found evidence for the molecular basis of gating the cloned K(ATP) by intracellular protons. Systematic constructions of chimerical Kir6.2-Kir1.1 channels indicated that full pH sensitivity required the N terminus, C terminus, and M2 region. Three amino acid residues were identified in these protein domains, which are Thr-71 in the N terminus, Cys-166 in the M2 region, and His-175 in the C terminus. Mutation of any of them to their counterpart residues in Kir1.1 was sufficient to completely eliminate the pH sensitivity. Creation of these residues rendered the mutant channels clear pH-dependent activation. Thus, critical players in gating K(ATP) by protons are demonstrated. The pH sensitivity enables the K(ATP) to regulate cell excitability in a number of physiological and pathophysiological conditions when pH is low but ATP concentration is normal.

Adenosine Triphosphate↗
PubMed2001

Studies directed towards the refinement of the pancratistatin cytotoxic pharmacophore.

Two deoxy-analogues of the anticancer/antiviral agent pancratistatin containing functionality complementary to the minimum structural pharmacophore were synthesized and subjected to anticancer screening. One of the analogues exhibited selective inhibition of certain tumor cell lines but was significantly less potent than the natural products. The minimum structural pharmacophore has now been refined from eight to three possible structures.

Amaryllidaceae Alkaloids↗