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Biomedical subjects

Tian Yang

Publications and source records attributed to Tian Yang.

15 recordsLinked to original sources

Functions of tandem-repeat galectins and domain coordination governs galectin-4 activity in grass carp (Ctenopharyngodon idella).

Galectins are β-galactoside-binding lectins that play essential roles in innate immunity. Among them, tandem-repeat galectins (TrGals), typically composed of two distinct carbohydrate-recognition domains (CRDs) connected by a linker peptide, are well established as key regulators of pathogen recognition and host defense in mammals. However, their structural diversity and immunological functions in teleost fish remain poorly understood. In this study, five TrGals (Gal-4, Gal-8a, Gal-8b, Gal-9, and Gal-9like) were identified in grass carp. Sequence and structural analysis revealed that Gal-8a/b, Gal-9, and Gal-9like possess the canonical two-CRD architecture, whereas Gal-4 uniquely contains four highly similar tandem-repeat domains. All five TrGals were broadly expressed across examined tissues, with predominant expression in the liver. Upon Aeromonas hydrophila infection, Gal-4, Gal-8a, Gal-8b, and Gal-9 were rapidly up-regulated at early time points (3-6 h). To elucidate the functional significance of CRD number, recombinant full-length CiGal-4 (CiGal4-full) and three truncated variants containing one, two, or three CRDs (CiGal4-1CRD, CiGal4-2CRD, and CiGal4-3CRD) were generated and systematically characterized. All recombinant proteins contained the conserved β-sheet structure typical of galectin CRDs. Functional assays revealed that CiGal4-full displayed the strongest growth-inhibitory activity against all tested bacteria, whereas CiGal4-1CRD showed the weakest effect. Notably, CiGal4-2CRD exhibited the most potent bactericidal activity, surpassing the full-length protein, while CiGal4-3CRD showed no further enhancement. CiGal4-full and CiGal4-2CRD showed superior carbohydrate-binding activities compared with the other variants. Collectively, these results reveal that CRD copy number alone does not linearly determine galectin function. Instead, domain organization and conformational coordination are critical for optimizing antimicrobial activity. This study provides new insights into the structure-function relationships and evolutionary diversification of galectins in teleosts and highlights their potential as novel antimicrobial and immunomodulatory agents in aquaculture.

Animals↗

Rice transcription factor bHLH25 confers resistance to multiple diseases by sensing H2O2.

Hydrogen peroxide (H2O2) is a ubiquitous signal regulating many biological processes, including innate immunity, in all eukaryotes. However, it remains largely unknown that how transcription factors directly sense H2O2 in eukaryotes. Here, we report that rice basic/helix-loop-helix transcription factor bHLH25 directly senses H2O2 to confer resistance to multiple diseases caused by fungi or bacteria. Upon pathogen attack, rice plants increase the production of H2O2, which directly oxidizes bHLH25 at methionine 256 in the nucleus. Oxidized bHLH25 represses miR397b expression to activate lignin biosynthesis for plant cell wall reinforcement, preventing pathogens from penetrating plant cells. Lignin biosynthesis consumes H2O2 causing accumulation of non-oxidized bHLH25. Non-oxidized bHLH25 switches to promote the expression of Copalyl Diphosphate Synthase 2 (CPS2), which increases phytoalexin biosynthesis to inhibit expansion of pathogens that escape into plants. This oxidization/non-oxidation status change of bHLH25 allows plants to maintain H2O2, lignin and phytoalexin at optimized levels to effectively fight against pathogens and prevents these three molecules from over-accumulation that harms plants. Thus, our discovery reveals a novel mechanism by which a single protein promotes two independent defense pathways against pathogens. Importantly, the bHLH25 orthologues from available plant genomes all contain a conserved M256-like methionine suggesting the broad existence of this mechanism in the plant kingdom. Moreover, this Met-oxidation mechanism may also be employed by other eukaryotic transcription factors to sense H2O2 to change functions.

Hydrogen Peroxide↗

Bulge cells of human hair follicles: segregation, cultivation and properties.

The bulge region of hair follicle has been reported as a putative reservoir of hair follicle stem cell (HFSC) for years; however, few studies were done about the characteristics of bulge-originated cells in vitro up to now. In this experiment, the bulge cells isolated from human hair follicles by enzymatic digestion and microdissection were cultured and passaged, and the morphological and biological features of cultured bulge cells were investigated by microscopy and immunocytochemistry. The result showed that new-proliferated cells could be observed on the second day after inoculation, and the quantity of the cells with a greater proliferation potential, reached a peak at the 6th day and maintained this higher level for several days. The mitotic figures of bulge cells were seen and these cells showed undifferentiated morphologic features. The bulge cells strongly expressed K19 and beta1-integrin, which are the markers of HFSC, in a descensive way with the culture time. The result indicates that the cultured bulge cell from human hair follicle possesses the properties of primitive cells and supports the hypothesis that HFSC resides in the bulge area.

Cell Differentiation↗

Patterns of nestin expression in human skin.

Nestin, a member of the sixth class of intermediate filament proteins, has long been known not only as a specific marker for central nervous system stem cells, but also as an indicator of the degree of neural stem cell differentiation. Immunohistochemical analysis showed nestin expression in the epidermis and the upper two-third of the hair follicle in non-balding human scalp skin, but not in the lower third of the follicle. Expression was very weak in the sebaceous gland. The level of nestin mRNA transcription was much higher in the bulge region of the follicle than in the sebaceous gland. These results show that the pattern of nestin expression is related to the differentiation of epidermal stem cells.

Epidermal Cells↗

Molecular cloning and identification of mouse Cklfsf2a and Cklfsf2b, two homologues of human CKLFSF2.

Human chemokine-like factor superfamily (CKLFSF) is a novel gene family comprising CKLF and CKLFSF1-8. Among them, CKLFSF2 is highly expressed in testis and may play important roles in male reproduction. Besides, it is very active during evolution and has two counterparts in mouse. For further study, we cloned the two mouse genes by EST assembly and RT-PCR methods and designated them as mouse Cklfsf2a and Cklfsf2b. Their predicted open-reading frames (ORFs) that encode 169 and 210 amino acids, respectively, were obtained; and their predicted full-length molecular sizes that are approximately 1.2 kb for mCklfsf2a and 0.9 kb for mCklfsf2b were confirmed by Northern blot analysis. Mouse Cklfsf2a and Cklfsf2b show similarities with human CKLFSF2 in the expression patterns that are abundant in testis, hematopoietic and immune tissues; as well as in the chromosome localizations that neighbor CKLFSF1 and 3. Their putative protein products have 47.6 and 45.5% identities with hCKLFSF2, respectively; both of them contain four potential transmembrane regions and MARVEL domains, which are also similar with hCKLFSF2. Functionally, they all can affect the transcriptional activity of androgen receptor in PC-3 and HeLa cells, but mCklfsf2a is a repressor while mCklfsf2b and hCKLFSF2 are enhancers. Taken together, we conclude that mouse Cklfsf2a and Cklfsf2b are two homologues of human CKLFSF2. Studies on them would provide much help in further investigation of the latter.

Amino Acid Sequence↗

Effects of intestinal trefoil, factor on colonic mucosa in experimental colitis of rats.

In order to investigate the protective effects of intestinal trefoil factor (ITF) on colonic mucosa in experimental colitis of rats, ITF was detected by RT-PCR and immunohistochemistry at different time points. Three days after colitis induction, rats were treated with either 0.9% saline solution or rhITF. Pathological changes and the expression of iNOS mRNA, NO, MDA and SOD were measured respectively. It was found that ITF was mainly located in goblet cells, significantly higher in model group than in normal group (P<0.05). rhITF could increase the iNOS mRNA expression and NO contents, and there was statistically significant difference between rhITF group and model group (P<0.05). rhITF also caused an increase of MDA and a decrease of SOD, but there was no significant difference between two groups. These results indicated that ITF has apparent therapeutic effects in ulcerative colitis, which may be associated with iNOS and NO.

Acetic Acid↗

Effects of exogenous wild-type P16 gene transfection on the expression of cell cycle-related proteins in bladder cancer cell line.

Cell cycle regulatory proteins are important indicators in determining progression through the cell cycle. Recent experimental evidence shows that active Cdk4-Cyclin D1 complexes help cells to pass through the R point, a point of no return, after which the cells become committed to a new round of replication. It is widely known that P16INK4a can arrest cells in the G1 phase, but how the expression of exogenous P16INK4 gene can affect the activity of Cdk4-Cyclin D1 remains unclear. In this study, using exogenous wild-type P16 gene, antibodies for P16, Cdk4, Cyclin D1 and Rb proteins, and primers for these genes, we examined the expression of exogenous wild-type P16 gene and the changes of cell cycle regulatory genes (Cdk4, Cyclin D1, and Rb) in human bladder cancer cells. The cell cycle analysis revealed that the proliferation of P16 gene-transfected cancer cells was inhibited after the transfection of exogenous wild P16 gene. The immunocytochemical results indicated that after the transfection of exogenous wild-type P16 gene, the expression of Cdk4, Cyclin D1, and Rb were negative in the nuclei, whereas the expression of P16 significantly increased in the nuclei and the cytoplasm. The RT-PCR results showed that the transcription of P16 gene increased significantly after the transfection, whereas the transcription of Cdk4, Cyclin D1, and Rb decreased. Our results suggest that the transfection of exogenous wild P16 gene induces the bladder cancer cells arrested in G0/G1 phase, and the increasing expression of P16 inhibits the expression of Cdk4, Cyclin D1, and Rb in nuclei. As a consequence, exogenous P16 has negative effects on the malignant proliferation of the bladder cancer cells, and it may be considered as target for potential anticancer drugs.

Cell Cycle↗

Cyclooxygenase-2 parallels invasive depth and increased MVD in transitional cell carcinoma.

To investigate the relationship between the expression of cyclooxygenase-2 (COX-2), and invasive depth and angiogenesis in human transitional cell carcinoma (TCC), the expression of COX-2 and von Willebrand factor (vWF) was detected in paraffin-embedded transitional cell carcinoma specimens from 50 patients with histologically confirmed bladder transitional cell carcinoma. Microvessels were counted and microvessel density (MVD) was calculated. The correlation between COX-2 and invasive depth, MVD and invasive depth, COX-2 and MVD were analyzed. COX-2 expression was significantly related to the invasive depth of tumors; it solely expressed in invasive TCC. The ratio of COX-2 positively stained samples in each stage correlated to the invasive stages; so did the proportion of COX-2 (+) cells in whole tumor cells. Also, MVD increased with the invasive stages of tumors and positively correlated to the invasive depth. The COX-2-positive TCC samples were highly vascular, their microvessel density was significantly higher than that of the COX-2-negative TCC samples. There was a positive correlation between MVD and COX-2 (+) cells proportion. Our findings suggested that COX-2 was positively correlated with the invasive depth and involved in the process of angiogenesis in human transitional cell carcinoma.

Carcinoma, Transitional Cell↗

Changes of uPA and uPA-R expression in the cornea after alkali burn.

PURPOSE: To investigate the expression levels of uPA and uPA-R in corneal repair after alkali burn. METHODS: The corneal alkali burn models were established in vitro and in vivo, then immunocytochemistry (ICC) of uPA/uPA-R and image analysis and statistical evaluation were performed to determine their expression levels both in vitro and in vivo. RESULTS: Compared with control group, the expressions of uPA and uPA-R after alkali burn had no significant increases till 6h, then increased rapidly from 12h to 24h and reached their maxima at 24h. From 24h, their expression levels decreased rapidly. In vivo, they rebounded again after 48h and attained their second peaks at 96h, respectively. After that, their expressions decreased again. The uPA-positive reaction mainly distributed in the cytoplasm while that of uPA-R mostly distributed on the cellular membrane. Their expression changes were similar to each other, both in vivo and in vitro. In vivo, uPA and uPA-R expressed and gathered in the basal layer of corneal epithelium. CONCLUSIONS: These results suggest that the time phase of 24h after wound is the typical stage associating with the expressional maximum of uPA and uPA-R both in vivo and in vitro. Furthermore, the results also imply that the expression changes of uPA and uPA-R correlate to the wound healing after the corneal alkali burn, uPA and uPA-R coordinate with each other to stimulate the wound healing.

Animals↗

Plasminogen activator inhibitor 2: expression and role in differentiation of epidermal keratinocyte.

Plasminogen activator inhibitor 2 (PAI-2) is an enzyme inhibitor which is involved in cell differentiation, tissue growth and regeneration. In this study, immunocytochemistry, in situ hybridization and confocal laser scanning microscopy were used to investigate the expression and role of PAI-2 in differentiation of keratinocytes in vitro. The result showed that in the mono-layer differentiated keratinocytes induced by high calcium concentration, the expression of PAI-2 and its mRNA increased significantly, accompanied by expression increase of the differentiation marker keratin 10; and in the multi-layer differentiated keratinocytes induced by high calcium, PAI-2 expressed strongly mainly in the keratinocytes of middle as well as upper stratified layers, while K10 expressed in the keratinocytes of all stratified layers. Furthermore, the changes of the parameters related to keratinocyte differentiation were detected after inhibition of PAI-2 functions by its antibody, and the data showed that when treated by PAI-2 antibody, involucrin in the keratinocytes envelope expressed increasingly with an altering distribution from part to the whole envelope area. Our results indicate that during differentiation of epidermal keratinocyte, PAI-2 expresses mainly in the more differentiated keratinocytes and may protect the terminal differentiated keratinocytes from prematuration through inhibiting involucrin expression in cornified envelope.

Animals↗

Urokinase plasminogen activator (uPA) is a positive regulator of outer root sheath keratinocyte proliferation.

Urokinase plasminogen activator (uPA), a serine proteinase, is important in the development and epidermal wound healing, and seems to play a regulatory role in the proliferation of mouse epidermal keratinocytes (KC). In the present study, we found detectable uPA expression in outer root sheath (ORS) KC in the early anagen phase in mouse vibrissa follicles, but not in the late anagen or in the telogen and categen phases. uPA was also detected in ORS KC cultured from neonatal mice vibrissa. Specific exogenous inhibitors of uPA, amiloride and uPA antibody, significantly reduced the proliferation of ORS KC. Thus uPA is consistently elevated in the hyperproliferative hair follicle KC, and inhibition of the enzyme decreases hair follicle KC proliferation. We deduce that uPA is a very important mediator of the hair follicle cycle because its activity correlates with ORS KC proliferation.

Animals↗

[Preparation, identification, and analysis on tissue chips of polyclonal anti-peptide antibody to chemokine-like factor 1].

OBJECTIVE: To prepare the polyclonal anti-peptide antibody against chemokine-like factor1 (CKLF1) and apply it to the expression and functional studies of CKLF1. METHODS: CKLF1 was analyzed with bioinformatics methods. The 16 amino acids sequence peptide was selected from CKLF1 C terminal end. Antibody was raised by immunizing rabbits with the peptide conjugated to keyhole limpet hemocyanin (KLH). RESULTS: A high titer polycolonal antibody was obtained from the rabbit against the peptide. ELISA analysis proved that the titer of rabbit serum against anti-peptide of CKLF1 was up to 10(-4). Western blot analysis revealed that it could react not only with recombinant CKLF1 expressed in a cell-Free Protein Biosynthesis System and Drosophila S2 cells, but also recognize the endogenous CKLFs in the tissue array. Positive staining was detected in the normal bronchial cartilage, gastric mucosa, and gastric smooth muscle tissues. Normal rectum and well-differentiated rectal carcinoma showed strong positive staining, but the poor-differentiated rectal carcinoma samples revealed negative staining. CONCLUSION: The anti-peptide antibody can specifically recognize CKLFs and may be a useful reagent for the detection of CKLF1.

Animals↗

Identification of eight genes encoding chemokine-like factor superfamily members 1-8 (CKLFSF1-8) by in silico cloning and experimental validation.

TM4SF11 is only 102 kb from the chemokine gene cluster composed of SCYA22, SCYD1, and SCYA17 on chromosome 16q13. CKLF maps on chromosome 16q22. CKLFs have some characteristics associated with the CCL22/MDC, CX3CL1/fractalkine, CCL17/TARC, and TM4SF proteins. Bioinformatics based on CKLF2 cDNA and protein sequences in combination with experimental validation identified eight novel genes designated chemokine-like factor superfamily members 1-8 (CKLFSF1-8). CKLFSF1-8 form gene clusters; the sequence identities between CKLF2 and CKLFSF1-8 are from 12.5 to 39.7%. Most of the CKLFSFs have alternative RNA splicing forms. CKLFSF1 has a CC motif and higher sequence similarity with chemokines than with any of the other CKLFSFs. CKLFSF8 shares 39.3% amino acid identity with TM4SF11. CKLFSF1 links the CKLFSF family with chemokines, and CKLFSF8 links it with TM4SF. The characteristics of CKLFSF2-7 are intermediate between CKLFSF1 and CKLFSF8. This indicates that CKLFSF represents a novel gene family between the SCY and the TM4SF gene families.

Alternative Splicing↗

Cyclooxygenase-2 increased the angiogenic and metastatic potential of tumor cells.

It seems certain that COX-2 is related to tumor and some data suggested that COX-2 might have relation to tumor malignance and angiogenesis. In order to elucidate the relationship between COX-2 and tumor invasive and angiogenic ability, we transfected human transitional cell carcinoma (TCC) cell line, EJ, permanently with a COX-2 expression vector or the mock vector. The EJ-COX(2) cells, which overexpressed COX-2, acquired increased invasiveness and angiogenic ability by activation of VEGF, uPA, and MMP-2. Increased invasiveness and angiogenic ability were reversed by treatment with either selective COX-2 inhibitor, NS-398, or dual COX inhibitor, indomethacin. These results demonstrate that overexpression of COX-2 can lead to phenotypic changes that alter the metastatic and angiogenic potential of TCC cancer cells.

Carcinoma, Transitional Cell↗