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M Li

Publications and source records attributed to M Li.

At least 595 records · Page 33Linked to original sources
PubMed1998

Ecotropic C-type retrovirus of B16 melanoma and malignant transformation of normal melanocytes.

We reported previously that B16,JB/RH and JB/MS melanomas of C57BL/6 mice express the common melanoma-associated antigen (MAA) recognized by MM2-9B6 monoclonal antibody (MAb). This MAA is encoded by the env gene of an ecotropic MuLV-type retrovirus that somatically emerged in melanomas of C57BL/6 mice. The potential role of this melanoma-associated retrovirus (MelARV) in melanoma formation remains unknown and has not been previously investigated. To test this, normal melanocyte lines (melan-a and C57M) of C57BL/6 mice were infected with the MelARV produced by B16BL6 melanoma. Infection of these melanocytes with the MelARV was associated with the appearance of the MAA recognized by MM2-9B6 MAb. Most of the infected melanocyte sublines were able to grow only in the presence of 12-O-tetradecanoylphorbol-13-acetate (TPA). Two infected melanocyte sublines showed morphological changes, were able to grow in the absence of TPA and, after inoculation into C57BL/6 mice, produced rapidly growing, highly pigmented tumors. These new melanomas, derived from the MelARV-infected melan-a and C57M melanocytes, were termed Meli-A1 and Meli-BL, respectively. Southern blot analysis of EcoRI- and HindIII-digested DNAs from these melanomas showed several retroviral insertion sites. One copy of MeIARV was found to be inserted at the end of the 6th leucine domain of the c-maf proto-oncogene, which encodes a basic region/leucine zipper transcription factor related to the AP-1 family that is able to form homodimers or heterodimers with Fos and Jun transcription factors. Our data indicate that c-maf is a common insertion site of MelARV in BL6, Meli-A1 and Meli-BL melanomas, whereas no such insertion site was found in the melanocytes infected with MelARV but not malignantly transformed. Thus, our data imply that the ecotropic MelARV that somatically emerged in B16 and other melanomas of C57BL/6 mice may play a role in malignant transformation.

Animals↗
PubMed1998

Peptide binding consensus of the NHE-RF-PDZ1 domain matches the C-terminal sequence of cystic fibrosis transmembrane conductance regulator (CFTR).

The Na+-H+ exchanger regulatory factor (NHE-RF) is a cytoplasmic phosphoprotein that was first found to be involved in protein kinase A mediated regulation of ion transport. NHE-RF contains two distinct protein interaction PDZ domains: NHE-RF-PDZ1 and NHE-RF-PDZ2. However, their binding partners are currently unknown. Because PDZ domains usually bind to specific short linear C-terminal sequences, we have carried out affinity selection of random peptides for specific sequences that interact with the NHE-RF PDZ domains and found that NHE-RF-PDZ1 is capable of binding to the CFTR C-terminus. The specific and tight association suggests a potential regulatory role of NHE-RF in cystic fibrosis transmembrane conductance regulator (CFTR) function.

Amino Acid Sequence↗
PubMed1998

Analysis of the DNA-binding site for Xenopus glucocorticoid receptor accessory factor. Critical nucleotides for binding specificity in vitro and for amplification of steroid-induced fibrinogen gene transcription.

In addition to the glucocorticoid receptor, DNA-binding proteins called accessory factors play a role in hormone activation of many glucocorticoid-responsive genes. Hormonal regulation of the gamma-fibrinogen subunit gene from the frog Xenopus laevis requires a novel DNA sequence that binds a liver nuclear protein called Xenopus glucocorticoid receptor accessory factor (XGRAF). Here we demonstrate that the recognition site for XGRAF encompasses GAGTTAA at positions -175 to -169 relative to the start site of transcription. This sequence is not closely related to the binding sites for known transcription factors. The two guanosines make close contact with XGRAF, as shown by the methylation interference assay. Single-point mutagenesis of every nucleotide in the 9-base pair region from positions -177 to -169 showed an excellent correlation between ability to bind XGRAF in vitro and ability to amplify hormone-induced transcription from DNA transfected into Xenopus primary hepatocytes. Conversely, XGRAF had little or no effect on basal transcription of the gamma-fibrinogen gene. Maximal hormonal induction also requires three half-glucocorticoid response elements (half-GREs) homologous to the downstream half of the consensus GRE. Interestingly, the XGRAF-binding site is immediately adjacent to the most important half-GRE. This close proximity suggests a new mechanism for activation of a gene lacking a conventional full GRE.

Animals↗
PubMed1998

Two distantly positioned PDZ domains mediate multivalent INAD-phospholipase C interactions essential for G protein-coupled signaling.

Drosophila INAD, which contains five tandem protein interaction PDZ domains, plays an important role in the G protein-coupled visual signal transduction. Mutations in InaD alleles display mislocalization of signaling molecules of phototransduction which include the essential effector, phospholipase C-beta (PLC-beta), which is also known as NORPA. The molecular and biochemical details of this functional link are unknown. We report that INAD directly binds to NORPA via two terminally positioned PDZ1 and PDZ5 domains. PDZ1 binds to the C-terminus of NORPA, while PDZ5 binds to an internal region overlapping with the G box-homology region (a putative G protein-interacting site). The NORPA proteins lacking binding sites, which display normal basal PLC activity, can no longer associate with INAD in vivo. These truncations cause significant reduction of NORPA protein expression in rhabdomeres and severe defects in phototransduction. Thus, the two terminal PDZ domains of INAD, through intermolecular and/or intramolecular interactions, are brought into proximity in vivo. Such domain organization allows for the multivalent INAD-NORPA interactions which are essential for G protein-coupled phototransduction.

Amino Acid Sequence↗
PubMed1998

Studying development of disease through temporally controlled gene expression in the salivary gland.

Multistep tumorigenesis proceeds through activation of oncogenes and inactivation of tumor suppressor genes. Initiating oncoproteins induce secondary changes that maintain transformation in the absence of original stimuli. Time-dependent reversal of SV40 T antigen (TAg)-induced hyperplasia was studied using temporally controlled gene expression. Targeting TAg expression to the submandibular salivary gland of transgenic mice produces focal hyperplasias at age two weeks, which extend through large areas of the gland by four months. At twelve months, fibrosis and tumor foci accompany hyperplasia. Hyperplasia reverses when TAg expression in discontinued at four months but not at seven months. Secondary changes that maintain transformation appear to be time dependent. The system can be used to identify genetic events resulting in phenotypic reversal at four months and to expose factors preventing its occurrence at seven months. Expression of other proteins can be targeted to the salivary gland, and temporally controlled gene deletions can also be made using this system.

Animals↗
PubMed1998

T-->G or T-->A mutation introduced in the branchpoint consensus sequence of intron 4 of lecithin:cholesterol acyltransferase (LCAT) gene: intron retention causing LCAT deficiency.

Previous mutations associated with lecithin:cholesteryl acyltransferase (LCAT) deficiency syndromes have been identified in the coding regions of the LCAT gene. However, recently, an intron mutation was found in a family in which three sisters presented with fish-eye disease (FED). The probands were shown to be heterozygotes for a mutation in intron 4. The respective T-->C nucleotide substitution, 22 bases upstream of the 3'-splice site, causes a null allele as the result of complete intron retention. Since the natural mutation occurs in a putative branchpoint consensus sequence of the intron, it was hypothesized that the point mutation may disrupt the splicing of the pre-mRNA. To further study the functional significance of the above thymine residue in the branchpoint sequence, we introduced other nucleotides at this position, i.e., LCAT Int-4 MUT-1 (T-->G) and LCAT Int-4 MUT-2 (T-->A). After stable transfection of the mutated pNUT-LCAT minigenes into BHK cells, we could detect neither LCAT activity nor LCAT protein in the culture medium of the pNUT-LCAT Int-4 MUT-1 and pNUT-LCAT Int-4 MUT-2 cell lines, as was previously described for the natural mutation. To determine the effects of the introduced mutations on pre-mRNA splicing, total RNA from transfected BHK cells was used for RT-PCR analysis. All BHK cell lines were shown to transcribe the integrated LCAT minigenes. However, the sizes of these LCAT messengers indicated that intron 4 was retained in the pNUT-LCAT Int-4 MUT-1 and pNUT-LCAT Int-4 MUT-2 cell lines. Subsequent sequence analysis of the RT-PCR products demonstrated that the unspliced intronic sequences contained the introduced mutations. In conclusion, the observed retention of intron 4 of the LCAT gene is the result of the specific loss of a thymine residue two bases upstream of the branchpoint adenosine residue in the putative branchpoint consensus sequence. The results confirm that a single base change in the branchpoint consensus sequence of an intron can cause human disease although this sequence is poorly conserved in mammals.

Animals↗
PubMed1998

Liposomal transfection efficiency and toxicity on glioma cell lines: in vitro and in vivo studies.

To evaluate the influence of cell type and cationic liposomal formulation on gene transfection efficiency three liposomes (lipofectin, lipofectamine and DOTAP) were used to transfect the human (A172 and MOG-G-CCM) and rodent (C6 and A15A5) glioma cell lines with the Lac Z gene. Parallel studies evaluated in vitro cytotoxicity and the neuropathological changes following intracerebral injection. The transfection efficiency and cytotoxicity of the liposomes varied both quantitatively and qualitatively between the cell lines. There were no behavioural disturbances following intrastriatal or hippocampal injection and the neuropathological changes at the injection sites were focal and similar for all liposomes. The influence of glial cell lineage on both liposomal transfection facility and cytotoxicity may have important implications for in vivo gene transfection in the central nervous system.

Animals↗
PubMed1998

Effect of melittin on PD, resistance and short-circuit current in the frog gastric mucosa.

In an in-vitro preparation of gastric mucosae of Rana pipiens, the effect of adding melittin to a concentration of 5x10-6 M in the secretory solution on the transepithelial potential difference (PD), resistance (R) and short-circuit current (Isc) was studied. In 20 min, melittin decreased the PD by 9.3 mV and R by 148 ohm cm2. These changes can be explained by a decrease in the resistance, RP, of the paracellular pathway. To determine whether specific-ion pathways were responsible for the decrease in R, the effect of melittin on the partial conductances of Cl-, K+ and Na+ was also studied using the ion substitution method. Melittin decreased the PD response to changes in nutrient Na+, K+ and Cl- and the PD response to changes in secretory Cl-, but did not affect PD responses to changes in secretory Na+ or K+. Therefore, melittin decreased the nutrient membrane partial conductances of Cl-, K+ and Na+ and secretory membrane partial conductance of Cl-, without affecting the secretory partial conductances of Na+ or K+. Initially, melittin increased Isc in regular and Cl--free but not in Na+-free solutions. There was a delayed decrease in Isc. The results indicate that melittin decreases RP, increases the Na+ conductance of the secretory membrane and inhibits, eventually, the Na+/K+-ATPase pump.

Animals↗
PubMed1998

Role of antioxidant enzymes on ionizing radiation resistance.

Exposure of cells to ionizing radiation leads to formation of reactive oxygen species (ROS) that are associated with radiation-induced cytotoxicity. The antioxidant enzymes manganese superoxide dismutase (MnSOD), copper-zinc superoxide dismutase (CuZnSOD), and glutathione peroxidase (GPx), are key intracellular antioxidants in the metabolism of ROS. We investigated the potential role of the three antioxidant enzymes in radioresistance by analyzing cell morphology, cell viability, and cellular radioresistance in stably transfected CHO cell lines overexpressing MnSOD, CuZnSOD, and GPx, respectively. We found that overexpression of human MnSOD substantially protected cells from ionizing radiation injury. Relative to MnSOD, GPx could slightly reduce the radiation sensitivity while the radioresistance in CuZnSOD expressing cells did not change significantly. The results suggested that MnSOD may play a central role in protecting cells against reactive oxygen species injury during ionizing radiation exposure among MnSOD, CuZnSOD, and GPx.

Animals↗
PubMed1998

Distinct functional stoichiometry of potassium channel beta subunits.

Shaker-type potassium channels play important roles in determining the electrical excitability of cells. The native channel complex is thought to be formed by four pore-forming alpha subunits that provide four interaction sites for auxiliary modulatory Kvbeta subunits. Because Kvbeta subunits possess diverse modulatory activities including either up-regulation or down-regulation of potassium currents, differential assembly of the alpha-beta complex could give rise to diverse current properties. However, the detailed physical and functional stoichiometry of the alpha-beta complex remains unknown. Kvbeta1 subunits reduce potassium currents through inactivation, whereas Kvbeta2 subunits enhance potassium currents by inhibiting the Kvbeta1-mediated inactivation and at the same time by promoting the surface expression of certain potassium channels. In this report we show that Kvbeta1 and Kvbeta2 of the Shaker-type potassium channels display distinct functional stoichiometry to interact with the Kv1 alpha subunits, a subfamily of Shaker-type potassium channels. The interaction of Kvbeta1 subunits with alpha subunits is consistent with the alpha4betan model, where n equals 0, 1, 2, 3, or 4, depending upon the relative concentration of alpha and beta subunits. The alpha4betan stoichiometry allows for gradual changes of the Kvbeta1-mediated inactivation. In contrast, Kvbeta2 subunits self-associate to form oligomers and interact with the alpha subunits via alpha4beta4 stoichiometry, which permits effective multivalent associations with alpha subunits. Such distinct functional stoichiometry of Kvbeta1 and Kvbeta2 provides a molecular mechanism that is well suited to their contrasting activities of up-regulation or down-regulation of potassium currents.

Animals↗
PubMed1998

[The relationship between p53 mutation and esophageal cancer biological behavior].

OBJECTIVE: To investigate the relationship between p53 mutation and esophageal cancer biological behavior as well as its prognostic role in esophageal cancer. METHOD: Mutations in exon 5-8 of p53 were screened in 30 sporadic esophageal cancers by a combination of PCR-SSCP and PCR-direct DNA silver sequencing. RESULTS: Mutations were found in 11 cases (36.7%). There were 9 point mutations, including 4 missense mutations, 2 nonsense mutations, 3 silent mutations. The other 2 cases were frameshift mutations due to base insertion and deletion. With statistical analysis, the p53 mutation rate was 56.3% in moderately and poorly-differentiated esophageal cancers, 14.3% in well-differentiated cases, with a highly significant difference between the two groups (P=0.025). The mutation rate of the cancers with all layers invaded (52.6%) was remarkably higher than that of the cancers without all layers invaded (9.1%)(P=0.024). A significantly higher rate of p53 mutation was also seen in cases with lymph nodes metastasis (61.5%) than in cases with no lymph nodes metastasis (17.6%) (P=0.024). CONCLUSION: The data suggest that there is a strong correlation between the presence of p53 mutation and the biological behavior of human esophageal cancer, such as histologic differentiation, invasion stage, and regional lymph nodes metastasis. Detection of p53 mutation can be helpful in identifying more progressive esophageal cancer and assessing prognosis. The study also discussed the dominant negative effect of p53 and the genetic effect of silent mutation.

Esophageal Neoplasms↗
PubMed1998

[Analysis of expression of FMR-1 gene in male patients with fragile X syndrome using RT-PCR].

OBJECTIVE: To search the relationship between fragile X male patients and expression of FMR-1 gene and to establish a rapid and simple method for screening male patients in general population. METHODS: Two typical fragile X families demonstrated by cytogenetics method were analyzed by using RT-PCR. RESULTS: No expression of FMR-1 gene in all male patients in the two families was noted, but all normal individuals showed expression of FMR-1 gene. There was an expression of HPRT gene as internal control in each of the individuals detected. CONCLUSION: There is a strong relationship between loss of expression of FMR-1 gene and clinical phenotype of the fragile X male patients. Further research of this method will introduce a more simple and rapid method for detecting male patients with fragile X syndrome. This method may be used for general population screening.

Fragile X Messenger Ribonucleoprotein 1↗
PubMed1998

Nuclear inclusions of the androgen receptor protein in spinal and bulbar muscular atrophy.

Spinal and bulbar muscular atrophy (SBMA) is an X-linked motor neuronopathy caused by the expansion of an unstable CAG repeat in the coding region of the androgen receptor (AR) gene. To study AR protein expression in normal and SBMA individuals, we used several antibodies that recognize AR protein, and analyzed neural and nonneural tissues by immunohistochemistry and western blotting. Both the normal and the mutant AR proteins were widely distributed, predominantly, but not exclusively, in the cytoplasm of neurons regardless of the pathological involvement, and predominantly in the nuclei of the nonneural tissues in both normal and SBMA individuals, with different expression levels of AR protein among different tissues. In the motor neurons of SBMA patients, there were AR-immunoreactive ubiquitinated nuclear inclusions that were detected by antibodies that recognize a small portion of the N terminus of the AR protein. Absence of other immunoreactive AR epitopes within the inclusion may be due to altered AR configuration, or masking of AR epitopes by other proteins, or proteolytic cleavage of the AR. Our data show that, in addition to the normal cellular distribution of the AR protein, mutant AR-bearing nuclear inclusions are present in SBMA.

Aged↗
PubMed1998

Hepatic enzyme induction and acute endocrine effects of 2,3,3',4',6-pentachlorobiphenyl in prepubertal female rats.

Polychlorinated biphenyls (PCBs) with the labile 2,3,6-substitution are important components of atmospheric and certain food chain exposures, but little is known about their biological activities. Chlorobiphenyl 110 (2,3,3',4',6-pentaCB) was investigated in weanling female rats dosed ip on days 21 and 22 and killed on day 23 of age. The initial preparation of CB 110 markedly induced 7-ethoxyresorufin O-dealkylase (EROD) activity and was found to be contaminated with coplanar 3,3',4,4',5-pentaCB (CB 126). The contaminated preparation (CB 110C) was purified with activated charcoal (CB 110P). The CB 110P induced pentoxyresorufin O-dealkylase (PROD), was weakly uterotropic and a modest depleter of serum thyroxine (T4). CB 110C caused increased liver weight, induced EROD, PROD, and UDP glucuronyl transferase activities and caused a greater depletion of serum T4; on the other hand, it suppressed the PROD induction and the uterotropic effect of CB 110P. Hepatic residues of CB 110 were a constant 2-3% of the dose while those of CB 126 (from CB 110C) increased with increasing dose to as much as 50% of the dose.

Animals↗
PubMed1998

Expression of low-affinity neurotrophin receptor p75NTR in the peripheral nervous system of human neuropathies.

Expression of low-affinity neurotrophin receptor (p75NTR) was immunohistochemically examined in the peripheral nerve trunks, dorsal root ganglia, sympathetic nerve ganglia and spinal cords in various human neurological diseases manifesting peripheral neuropathies. p75NTR was expressed in the nerves with axonal degeneration, and was also prominent in the nerves with newly regenerating axons. In contrast, axonal pathology tended to reduce the expression of p75NTR in the neuronal perikarya of the dorsal root ganglion and sympathetic nerve ganglion neurons. In the ventral and lateral horn cells, the p75NTR immunoreactivity was not detected in the normal and diseased nerves except for amyloid polyneuropathy. These p75NTR expressions in the diseased human peripheral nervous tissues would be regulated by an underlying pathology-related process, and could play a role in peripheral nerve repair.

Aged↗
PubMed1998

Nonneural nuclear inclusions of androgen receptor protein in spinal and bulbar muscular atrophy.

Spinal and bulbar muscular atrophy is an X-linked motor neuronopathy caused by the expansion of an unstable CAG repeat in the coding region of the androgen receptor (AR) gene. Nuclear inclusions of the mutant AR protein have been shown to occur in the spinal motor neurons of spinal and bulbar muscular atrophy (Li M, Kobayashi Y, Merry D, Tanaka F, Doyu M, Hashizume Y, Fischbeck KH, Sobue G: Nuclear inclusions in spinal and bulbar muscular atrophy. Ann Neurol 1998 (in press)). In this study, we demonstrate the tissue-specific distribution, immunochemical features, and fine structure of nuclear inclusions of spinal and bulbar muscular atrophy. Nuclear inclusions were observed in affected spinal and brainstem motor neurons, but not in other, nonaffected neural tissues. Similar nuclear inclusions occurred in nonneural tissues including scrotal skin, dermis, kidney, heart, and testis, but not in the spleen, liver, and muscle. These inclusions had similar epitope features detectable by antibodies that recognize a small portion of the N-terminus of the AR protein only, and they were ubiquitinated. Electron microscopic immunohistochemistry showed dense aggregates of AR-positive granular material without limiting membrane, both in the neural and nonneural inclusions. These findings indicate that nuclear inclusions of AR protein are present in selected nonneural tissues as well as in neurons that degenerate in spinal and bulbar muscular atrophy, suggesting that a common mechanism underlies in the formation of neural and nonneural nuclear inclusions.

Aged↗
PubMed1998

Tapping mode atomic force microscopy of hyaluronan: extended and intramolecularly interacting chains.

The extracellular matrix polysaccharide hyaluronan has been examined by tapping mode atomic force microscopy. High molecular weight hyaluronan was deposited on mica from dilute aqueous solution and imaged in air. Long unbranched chains could be observed and were found to be compatible with the known covalent structure of hyaluronan. In addition, chains with evidence of intramolecular association were observed. In the simplest cases, the association took the form of loops stabilized by antiparallel double-stranded (probably double-helical) segments. In other cases, the polarity of the associated regions could not be determined. Extensive intramolecular association in long hyaluronan chains resulted in a fenestrated structure of the same type as that formed by intermolecular association at higher concentrations.

Animals↗