Biomedical subjects

Jing An

Publications and source records attributed to Jing An.

30 records · Page 2Linked to original sources

PubMed2004

Hematoporphyrin monomethyl ether photodynamic damage on HeLa cells by means of reactive oxygen species production and cytosolic free calcium concentration elevation.

Hematoporphyrin monomethyl ether (HMME) is a novel and promising porphyrin-related photosensitizer for photodynamic therapy (PDT). HMME-PDT-induced cell death and its mechanisms were investigated in HeLa cells. We demonstrated that HMME-PDT could induce cell death through both necrosis and apoptosis. Sodium azide (the singlet oxygen quencher) or D-mannitol (the hydroxyl radical scavenger) could protect HeLa cells from the apoptosis and necrosis induced by HMME-PDT, showing that reactive oxygen species (ROS), such as singlet oxygen and hydroxyl radical, played a decisive role in HMME-PDT-induced HeLa cells death. Sodium azide or D-mannitol also inhibited HMME-PDT-mediated [Ca2+]i elevation. Cytochrome C (Cyto C) release from mitochondria into cytosol and Caspase-3 activation after HMME-PDT were inhibited by BAPTA/AM (an intracellular calcium chelator). These results demonstrated that ROS generated in HeLa cells by HMME-PDT-induced apoptosis may be through [Ca2+]i elevation which mediates Cyto C release and Caspase-3 activition and initiates the subsequent late stages of apoptosis.

Apoptosis↗

PubMed2004

Critical upstream signals of cytochrome C release induced by a novel Bcl-2 inhibitor.

Cytochrome c release is a central step in the apoptosis induced by many death stimuli. Bcl-2 plays a critical role in controlling this step. In this study, we investigated the upstream mechanism of cytochrome c release induced by ethyl 2-amino-6-bromo-4-(1-cyano-2-ethoxy-2-oxoethyl)-4H-chromene-3-carboxylate (HA14-1), a recently discovered small molecule inhibitor of Bcl-2. HA14-1 was found to induce cytochrome c release from the mitochondria of intact cells but not from isolated mitochondria. Cytochrome c release from isolated mitochondria requires the presence of both HA14-1 and exogenous Ca(2+). This suggests that both mitochondrial and extramitochondrial signals are important. In intact cells, treatment with HA14-1 caused Ca(2+) spike, change in mitochondrial membrane potential (Delta psi(m)) transition, Bax translocation, and reactive oxygen species (ROS) generation prior to cytochrome c release. Pretreatment with either EGTA acetoxymethyl ester or vitamin E resulted in a significant decrease in cytochrome c release and cell death induced by HA14-1. Furthermore pretreatment with RU-360, an inhibitor of the mitochondrial Ca(2+) uniporter, or with EGTA acetoxymethyl ester, but not with vitamin E, prevented the HA14-1-induced Delta psi(m) transition and Bax translocation. This suggests that ROS generation is an event that occurs after the Delta psi(m) transition and Bax translocation. Together these data demonstrate that the Ca(2+) spike, mitochondrial Bcl-2 presensitization, and subsequent Delta psi(m) transition, Bax translocation, and ROS generation are important upstream signals for cytochrome c release upon HA14-1 stimulation. The involvement of endoplasmic reticulum and mitochondrial signals suggests both organelles are crucial for HA14-1-induced apoptosis.

Apoptosis↗

PubMed2004

Dengue-specific CD8+ T cells have both protective and pathogenic roles in dengue virus infection.

To analyze roles of memory T cells in the pathogenesis of dengue (DEN) virus infection, a DEN virus-specific CD8+ cell clone (2D42 cell) was employed to investigate its in vivo function after DEN virus infection using an animal model. HepG2 grafted severe combined immunodeficient (HepG2-grafted SCID) mice were divided into three groups--group A: HepG2-grafted SCID mice were inoculated intraperitoneally (ip) with 2D42 cells and then ip-infected with DEN virus type 2 (DEN-2); group B: HepG2-grafted SCID mice were inoculated with naive mouse thymocytes (NMT) and then ip-infected with DEN-2; group C: HepG2-grafted SCID mice were ip-infected with DEN-2 alone. Eighty percentage of group A mice died at average day 12.8 post-infection (p.i.) and 20% of them recovered from the disease after showing clinical signs and survived more than 3 months. They showed severe manifestations including dramatically decreased platelet count, decreased hematocrit, anemia, viremia and high frequency of histopathological changes in several organs. All of group B mice also showed the above severe clinical signs. One hundred percentage mortality rate was noted in these mice and death occurred at average day 10.8 p.i., which was the earliest among three groups. Although the mice from group C showed 100% mortality rate and similar clinical signs, death observed in these mice occurred at average day 17.4 p.i. and the manifestations were slight and developed slowly. Our results suggested both protective and pathogenic roles for DEN-specific CD8+ T cell in DEN virus infection, whereas NMT did not provided any protection.

Animals↗

PubMed2004

[Electron microscopic observations of alveoli development and epithelial differentiation in the fetal lung of the Chinese].

OBJECTIVE: To observe the development of the alveoli and the differentiation of the epithelium in the fetal lung of Chinese. METHODS: Chinese human lung tissues were obtained from 20 fetuses of 10 to 34 weeks old, and the ultrastructural changes of the alveolar epithelial cells were observed using both optical and transmission electron microscope. RESULTS: At the first 10 to 16 weeks of development, the lung showed a tubular structure, the epithelial cells were columnar and poorly differentiated. At weeks 17 to 24, the bronchioles increased and the epithelial cells were differentiated to approach maturity. A few lamellar bodies were observed in the cuboidal lining cells with the majority being type II alveolar cells at 25 weeks, when the primitive alveolar was formed. In this stage many primitive epithelial cells could be seen, which appeared primordial in form before evolving into type II and type I cells and perhaps served as epithelial stem cells or progenitor cells. From about 26 weeks, type I cells were observed but most of the epithelial cells were type II cells. Type I cells began to increase in the alveolar epithelium until 32 weeks. CONCLUSIONS: The alveolar type II cells and type I cells are probably both derived from the same precursor cells or stem cells, and no evidence has shown that type II cells may differentiate into type I cells during the development of the fetal lung.

Cell Differentiation↗

PubMed2004

Expression of nestin, an intermediate filament protein, in human fetal hepatic stem cells.

OBJECTIVE: To explore a new oval cell marker by examining the expression of nestin in human fetal hepatic stem cells in various stages during human fetal development. METHODS: Human fetal tissue samples of various developmental stages were obtained and fixed for preparation of paraffin sections. The expression of nestin, generally recognized as a specific marker for neural precursor cells, in fetal hepatic stem cells was detected using immunocytochemical staining (ABC method). RESULTS: Nestin-positive oval cells, characterized to be hepatic stem cells, were found in the limiting plate, which arranged tightly in monolayer to form ductules that encapsulated the early-stage portal area in a sheath-like fashion, and also partially encapsulated the primary portal area. With the development of the secondary portal areas, the nestin-positive oval cells gradually centered around the Hering canal. In addition, some nestin-positive mononuclear cells were found scattering in the hepatic cords and sinusoids, especially in the mesenchymal tissue of the portal areas. CONCLUSION: Nestin-positive oval cells exist in human fetal liver and their origin and function await further investigation.

Fetus↗

PubMed2004

[Morphological studies on embryonic gonad development in Kunming mice].

OBJECTIVE: To observe the morphological changes of embryonic gonad in Kunming mice in different developmental stages. METHODS: HE stained paraffin sections of the embryonic mouse gonads in different developmental stages were prepared for evaluation of the morphological changes. RESULTS: The genital ridge was identified on the 11th day after the mating and undifferentiated gonad detected on the 12th day, while 1 d later, sexual identification was possible by examining the sections. The male gonad developed into testis by forming seminiferous tubules and Sertoli cells on the 14th day. On the 16th day, the female gonad evolved into the ovary with obvious differentiation of the cortex and medulla and till the 20th day, more seminiferous tubules occurred in the testis with cavities containing numerous spermatogenic cells at different developing phases; also observed was a mass of rough sex cords that produced ovarian follicles surrounding the ovary and contained large quantity of primitive oogonia. CONCLUSIONS: Days 11-12 after mating might be the best time for separating primordial germ cells from undifferentiated mouse gonad.

Animals↗

PubMed2004

[Expression of human tissue-type plasminogen activator in cow mammary gland].

OBJECTIVE: To construct an expression vector for highly efficient expression of tissue-type plasminogen activator (t-PA) confined in the mammary gland of cow to develop a cow mammary gland bioreactor. METHODS: RT-Touch down-PCR was employed to amplify human tissue-type plasminogen activator (t-PA) cDNA, which was digested with the restriction enzymes and subsequently cloned into the vector pSP72 for constructing specific fusion gene only expressed in the mammary gland. The fusion gene was then transferred into the mouse zygote and the mammary gland tissue of mice and cows. RESULTS: t-PA was detected in the milks of mice and cows after the transgenic manipulation with microinjection and mammary gland injection of the fusion gene. CONCLUSIONS: The vector we constructed can effectively induce t-PA expression in the mammary gland, which is not influenced by different transgenic methods. The expression level of t-PA, however, is significantly higher in the milk of cows than in the milk of mice, suggesting the species-specific difference in milk protein regulating system possibly is due to different factors and regulatory systems.

Animals↗

PubMed2004

[Expression of telomerase reverse transcriptase and its significance in human fetal lung development].

OBJECTIVE: To investigate the expression and distribution of telomerase reverse transcriptase (hTERT) and its significance during the development of epithelial stem cells in human fatal lungs. METHODS: Human lung tissues were obtained at abortion from 37 fetuses 10-34 weeks of gestational age with parental consent, and the expression of hTERT in these tissues was examined by immunohistochemistry. RESULTS: In fetuses of 10 weeks old, hTERT was detected mainly in the epithelial cells of the proximal bronchi in the fetal lung, and migrated gradually to the distal bronchi as the lung developed. The hTERT expression level peaked at 17-20 weeks of gestation and then decreased over the period of alveolarization that occurred at about 25-26 weeks. In the later gestation period, strongly positive epithelial cells became concentrated in discrete patches near the basal membrane of the airway and scattered in only a few of the pulmonary alveoli. Morphologically, these cells resembled the basal cells and type II alveolar epithelial cells (AEC) or their progenitor cells. CONCLUSIONS: Telomerase expression and activity are hallmarks of the pulmonary stem or progenitor cells that maintain undifferentiated state and self-renewal capacity, and it plays a crucial role in the normal differentiation and regeneration of the bronchial and alveolar epithelial cells to maintain the epithelial integrity.

DNA-Binding Proteins↗

PubMed2003

The pathogenesis of spinal cord involvement in dengue virus infection.

To investigate the mechanisms of dengue (DEN) virus transmission within the spinal cord, severe combined immunodeficient mice were intracerebrally inoculated with DEN virus type 2. After inoculation, a high virus titer and antigens were detected in the brain and spinal cord. At early stages of the infection, ultrastructural examinations showed that a few virions were present in the cytoplasm of ependymal cells lining the central canal. As the infection progressed, virions were observed in the lumen of the rough endoplasmic reticulum (RER), RER-derived vesicles and the Golgi region of infected neurons. These data suggest that the inoculated DEN virus might spread to the neurons of the spinal cord via the cerebral spinal fluid and cause several neuronal pathological responses. Moreover, DEN virus was also observed in myelinated and unmyelinated nerve fibers and typical neuronal synapses. Some virion-containing vesicles appeared to be fused with the membrane of presynapses, indicating that neuron-to-neuron transport of DEN virus might occur in the spinal cord. Additionally, anterior, lateral and posterior horns of the spinal cord exhibited different numbers of the positive neurons and different staining intensities of the DEN antigen during the infection. This difference likely represents variation of susceptibility to the DEN virus among the neurons of the spinal cord.

Animals↗

PubMed2003

Mutation analysis of the Smad3 gene in human osteoarthritis.

Osteoarthritis (OA) is the most common joint disease worldwide. Recent studies have shown that targeted disruption of Smad3 in mouse results in OA. To reveal the possible association between the Smad3 gene mutation and human OA, we employed polymerase chain reaction-single strand conformation polymorphism and sequencing to screen mutations in all nine exons of the Smad3 gene in 32 patients with knee OA and 50 patients with only bone fracture. A missense mutation of the Smad3 gene was found in one patient. The single base mutation located in the linker region of the SMAD3 protein was A --> T change in the position 2 of codon 197 and resulted in an asparagine to isoleucine amino-acid substitution. The expressions of matrix metalloproteinase 2 (MMP-2) and MMP-9 in sera of the patient carrying the mutation were higher than other OA patients and controls. This is the first report showing that the Smad3 gene mutations could be associated with the pathogenesis of human OA.

DNA Mutational Analysis↗

PubMed2003

[Isolation, identification and in vitro culture of human glomerular mesangial cells].

OBJECTIVE: To define the optimal condition to culture human glomerular mesangial cells in vitro. METHODS: With a three-layer micropore filter device, we separated the glomerular mesangial cells from normal human, with were subjected to identification with immunohistochemical method. Fluorescein isothiocyanate (FITC) staining was performed for labeling of the monoclonal antibodies so as to identify the cells' superficial markers including factor VIII, keratin, cytokeratin, desmine, et al. Glomerular endothelial cells isolated from the same human tissues were also cultured in vitro under the same condition, serving as negative control. RESULTS AND CONCLUSION: The antibodies related to anti-factor VIII, and those of antikeratin and anticytokeratin were negative in the isolated glomerular mesangial cells, while the antibodies for desmine, anti-collagen IV, anti-fibronectin, and anti-laminin were positive, offering evidence for confirming the identification of the isolated and cultured mesangial cells.

Cell Separation↗

PubMed2002

Exploring the stereochemistry of CXCR4-peptide recognition and inhibiting HIV-1 entry with D-peptides derived from chemokines.

Chemokine receptor CXCR4 plays an important role in the immune system and the cellular entry of human immunodeficiency virus type 1 (HIV-1). To probe the stereospecificity of the CXCR4-ligand interface, d-amino acid peptides derived from natural chemokines, viral macrophage inflammatory protein II (vMIP-II) and stromal cell-derived factor-1alpha (SDF-1alpha), were synthesized and found to compete with (125)I-SDF-1alpha and monoclonal antibody 12G5 binding to CXCR4 with potency and selectivity comparable with or higher than their l-peptide counterparts. This was surprising because of the profoundly different side chain topologies between d- and l-enantiomers, which circular dichroism spectroscopy showed adopt mirror image conformations. Further direct binding experiments using d-peptide labeled with fluorescein (designated as FAM-DV1) demonstrated that d- and l-peptides shared similar or at least overlapping binding site(s) on the CXCR4 receptor. Structure-activity analyses of related peptide analogs of mixed chiralities or containing alanine replacements revealed specific residues at the N-terminal half of the peptides as key binding determinants. Acting as CXCR4 antagonists and with much higher biological stability than l-counterparts, the d-peptides showed significant activity in inhibiting the replication of CXCR4-dependent HIV-1 strains. These results show the remarkable stereochemical flexibility of the CXCR4-peptide interface. Further studies to understand the mechanism of this unusual feature of the CXCR4 binding surface might aid the development of novel CXCR4-binding molecules like the d-peptides that have high affinity and stability.

Alanine↗