Biomedical subjects

Xia Wang

Publications and source records attributed to Xia Wang.

At least 91 records · Page 5Linked to original sources

PubMed2003

Alterations in mitochondria membrane potential and oxidative stress in infertile men: a prospective observational study.

OBJECTIVE: To evaluate the mitochondrial membrane potential (MMP) of spermatozoa and its correlation with semen parameters and production of reactive oxygen species (ROS) in infertile men and healthy donors. DESIGN: Controlled prospective study. SETTING: Male infertility clinic, Glickman Urological Institute, The Cleveland Clinic Foundation, Cleveland, Ohio. PATIENT(S): Nineteen infertile men and 7 healthy volunteers. INTERVENTION(S): Standard semen analysis, assessment of MMP and ROS production in spermatozoa. The MMP was assessed by flow cytometry using the probe carbocyanine DiOC(6)(3) and ROS was measured with chemiluminescence assay using luminol. MAIN OUTCOME MEASURE(S): The results of MMP are reported as the median interquartile range (IQR) number of cells counted in different areas of fluorescence. Results of ROS measurement are expressed as x10(6) counted photons per minute per 20 million sperm (cpm). RESULT(S): The patients with abnormal semen parameters had a significantly lower MMP [1337.7 (1066.38, 1879.2)], and higher ROS [1.12 (0.26, 3.86)] than the donors [MMP: 2482.9 (2162.5, 3520.6)] and [ROS: 0.10 (0.01, 0.14)]. The MMP was positively correlated with sperm concentration (r = 0.62) and negatively correlated with the ROS produced (r = -0.45). CONCLUSION(S): Measuring MMP in spermatozoa provides useful information about a man's fertility potential. Increased ROS production by spermatozoa is associated with a decreased MMP.

Carbocyanines↗

PubMed2003

[Screening of proteins interacting with tTG in HeLa cells].

Tissue transglutaminase(tTG) belongs to a class of transglutaminase family which is up-regulated in almost all cells apoptosis and is thought to be closely related to cell apoptosis. To investigate the mechanism of tTG in cell apoptosis, yeast two hybrid system was used to screen HeLa cDNA library. One of the 17 positive clones we have obtained encoded the glutamine-rich carboxyl terminus of TIAR, and this interaction between tTG and TIAR, which was finely regulated by Ca(2+), was proved in vitro by GST pull-down. These findings suggest that tTG might affect the function of TIAR by a calcium-dependent posttranslational modification and the interaction might possibly be involved in the regulation of cell apoptosis.

Amino Acid Sequence↗

PubMed2003

[Biological function of fusion protein ATF-PAI2CD].

To express the fusion protein ATF-PAI2CD (urokinase-type plasminogen activator amino terminal fragment-plasminogen activator inhibitor type 2 with the region inter C and D helices deleted ) gene in E.coli and determine the biological characterization of fusion protein ATF-PAI2CD, the cDNA fragment encoding ATF-PAI2CD was cloned into the expression vector pLY-4 and transformed into E.coli JF1125. After temperature induction, the expression amount of ATF-PAI2CD account for 15% of total bacterial protein. The result was confirmed by Western blot. ATF-PAI2CD protein was isolated and purified by washing and solubilization of inclusion body, renaturation and ion exchange chromatography. The final product displayed a single band with a corresponding molecular weight 62 kD in SDS-PAGE. The purity was over 90%, the protein yield was 50% and the specific activity was 12 000 IU/mg. The PAI activity was measured by chromogenic assay. The purified fusion protein inhibited urokinase-type plasminogen activator as measured by milk-agarose plate assay, and bound to human lung cancer cells via uPA receptor (uPAR), which was confirmed by radio competition experiments. The results indicate that the biological characteristics of ATF-PAI2CD were very similar to those of the wide type PAI-2 (or mutants PAI-2, PAI-2CD) and to pro-uPA in binding to uPAR-bearing cells.

Escherichia coli↗

PubMed2003

Identification of interaction between PAI-2 and IRF-3.

HeLa cells transfected with plasminogen activator inhibitor-2 ( PAI-2 ) were protected from TNF- alpha-induced apoptosis. The apoptosis protection by PAI-2 is dependent on a 33 amino acids fragment between helix C and D of PAI-2, which may be due to the interaction of PAI-2 with some intracellular proteins. In this study, the yeast two-hybrid system was used to screen a HeLa cells cDNA library constructed during apoptosis with the fragment between helix C and D of PAI-2 as bait. We retrieved a clone that encodes 98 amino acids of C-terminus of interferon regulatory factor-3 (IRF-3). Co-immunoprecipitation experiments confirmed the interaction between PAI-2 and IRF-3 in vivo. IRF-3 belongs to a family of the IRF transcription factors and has been shown to participate in a large number of biological processes. These data suggest that IRF-3 may be involved in the apoptosis protection and antiviral function of PAI-2.

Apoptosis↗

PubMed2003

[Leg re-contouring by using a new technique of partially removing gastrocnemius].

OBJECTIVE: To evaluate a method to partially remove gastrocnemius for improvement of the contour of the leg. METHODS: Thirteen patients with bulked calf were undergoing the treatment. With a self-designed apparatus, the gastrocnemius was partially removed by placing the device in the muscle through an incision in the popliteal fossa. The follow-ups were carried out for 1-6 months. RESULTS: Thirteen patients were successfully treated by above mentioned technique. The removed amount of muscle was weighted between 11 g and 201 g, averaged 77 g. All of the patients could be able to walk three days after the operation. But, the normal walk had to take 1 month after of the surgery for recovering. The function of the ankle joint was not obviously influenced. CONCLUSION: The above mentioned technique is a safe and effective method for re-contouring the leg.

Cosmetic Techniques↗

PubMed2003

[Internalization of antibody-targeted immunonanoparticles into human hepatoma cells and its reversal effect on MDR].

OBJECTIVE: To observe if antibody-targeted immunonanoparticles could internalize into sensitive and multidrug resistance (MDR) cells of human hepatoma; and to study if the immunonanoparticles could reverse the MDR. METHODS: Human hepatoma-specific adriamycin-loaded human serum albumin immunonanoparticles (HAb18-ADR-HSA-NP) were incubated with human hepatoma sensitive cell line (SMMC-7721) or MDR cell line (SMMC-7721/MDR+) and the internalization of immunonanoparticles were observed by laser confocus microscopy, scanning electron microscopy and transmission electron microscopy; MTT colorimetric assay was used for assaying in vitro cytotoxicities of HAb18-ADR-HSA-NP to the resistant variant cells. Then based on these data, IC50 value of the immunonanoparticles and RF (Resistant Factor) of MDR cells were calculated. RESULTS: Laser confocus microscopy showed that many fluorescent particles (labeled immunonanoparticles) tightly adsorbed to SMMC-7721 cells and were also seen in cytoplasm of SMMC-7721 cells. When incubated with immunonanoparticles at 37 degrees C, many of immunonanoparticles were visualized in cytoplasm of SMCC-7721 or SMCC-7721/MDR+. These immunonanoparticles-contained cells exhibited damaged ultrastructures and the damage degree depended on incubation time. When the human hepatoma cells were pretreated with HAb18 antibody and incubated with immunonanoparticles, few immunonanoparticles were seen in cytoplasm of the cells, suggesting antibody-specific internalization of the immunonanoparticles. Scanning electron microscopy demonstrated specific binding of the immunonanoparticles to the resistant variant cells. That immunonanoparticles exerted enhanced cytotoxicity to the resistant variant cells was demonstrated by a decrease of RF value of MDR cells, compared with free ADR (4.4 vs. 2.1). CONCLUSION: Human hepatoma-specific adriamycin-loaded human serum albumin immunonanoparticles could specifically internalize into sensitive or multidrug resistance cells of human hepatoma via antibody direction. The immunonanoparticles could enhance the sensitivity of MDR cells to ADR cytotoxicity, suggesting its reverse effect on MDR.

Antibiotics, Antineoplastic↗

PubMed2003

[The effect of anti-bovine serum albumin B cell clone in mice on preparation of McAb].

OBJECTIVE: To investigate the effect of anti-bovine serum B cell clone in mice on preparation of McAb. METHODS: Supernatant of hybridoma cell was double screened with rSIFN-co and BSA by indirect enzyme-linked immunosorbent assay. After BSA absorption, the inhibition effects on culture supernatant of double positive hybridoma cell clone and rSIFN-co single positive hybridoma cell clones and on normal mouse serum were investigated separately. RESULTS: One double positive cell clone and three rSIFN-co single positive clones were established. The reaction of rSIFN-co with the supernatant of the double positive clone was inhibited obviously on condition that the supernatant was absorbed with BSA in advance. CONCLUSION: In the screening of hybridoma cell, there is rSIFN-co and BSA double positive hybridoma cell clone, which suggests the possible existence of innate anti-BSA B cell clone in mice; they can cause false-positive reaction in the preparation of McAb. Thus, double screening with specific antigen and BSA is beneficial.

Animals↗

PubMed2003

[Study on post-laryngectomy partial laryngeal defect repaired with omohyoid myofascial flap].

OBJECTIVE: To explore the clinical effect on defect of post-laryngectomy partial laryngeal repaired with omohyoid myofascial flap. METHOD: Twenty-four cases of supraglottic and glottic cancer were treated surgically. The tumors and involved cartilages were removed entirely. The omohyoid myofascial flap was utilized to repair the defects of larynx and reconstruct the glottis. For the cases of unilateral fixation of arytenoid, the arytenoids were removed and replaced by a piece of omohyoideus linked to hyoid bone, then were covered with mucous. The cases above T2 supraglottic and T3 glottic received radiotherapy. RESULT: The following-up time is 1-5 years. One case of supraglottic cancer(T3) dies from local recurrence after operation for 2 years and 9 months. One case of glottic cancer(T3) dies from neck and bone metastasis after operation for 4 years and 3 months. Swallow function was preserved in all cases. The rate of deconnalation was 95.8%. 91.7% patients had laryngeal functions restored. CONCLUSION: For selected cases, using omohyoid myofascial flap to repair the partial laryngeal defect and reconstruct glottis is available and practicable.

Aged↗

PubMed2003

[Detection of serum heat shock protein 70 level in patients with hepatitis C infection and its role in cytotoxicity of CTLs induced by HSP70-HCV peptide complex].

AIM: To detect serum HSP70 in patients with hepatitis C and evaluate its role in cytotoxicity of specific CTLs induced by HSP70-HCV peptide complex. METHODS: The serum HSP70 level in patients infected with HCV and normal individuals were detected by ELISA. The relationship between anti-HCV antibody and HSP70 was evaluated. Peripheral blood mononuclear cells (PBMCs) were activated by HSP70-HCV peptide complex and then 4 hour (51)Cr release assay were used to detect the killer activity of CTLs. RESULTS: Detection rates of HSP70 were 82.1%(22/28) and 18.8% (9/48) in patients with anti-HCV antibody and in normal individuals, respectively. There was significant correlation between the detection rate of HSP70 and HCV infection (chi(2)=28.77, P<0.01). The level of serum HSP70 in patients infected with HCV was markedly more higher than that in normal individuals. CTLs from one patient induced by HSP70-HCV C region peptide (DLMGYIPAV) complex could lyse autologous B lymphoblastoid cell lines (BLcLs) with killer rate of 37.8%, whereas CTLs induced by peptide alone could not lyse autologous BLcLs cells. CONCLUSION: HCV infection can induce over expression of HSP70. HSP70 may have the action of enhancing presentation of HCV epitope peptide and promoting clearance cells infected with HCV.

HSP70 Heat-Shock Proteins↗

PubMed2002

[Differentiation of rat bone marrow stromal cells into neuron induced by baicalin].

OBJECTIVE: To study the possibility that induced marrow stromal cells (MSCs) to neuron in vitro by baicalin, a kind of flavonoid isolated from an important medicinal plant Scutellariae Radix. METHODS: MSCs from adult SD rats were induced by baicalin in serum-free medium for 6 days. The expression of neuronal or glia specific markers and gene was evaluated by indirect immunofluorescence cytochemistry staining, Western blot, and reverse transcriptase-polymerase chain reaction (RT-PCR). The percentage of apoptotic cells at 6 days after baicalin treatment was measured by terminal deoxynucleotidyl transferase- mediated dUTP-biotin nick end-labeling (TUNEL) assay. Moreover, differentiating cells that labeled by green fluorescent protein vector were transplanted to the striatum of adult SD rats after 6d of baicalin treatment. RESULTS: After induction for 6 d, MSCs displayed neuronal morphologies, such as pyramidal cell bodies and processes formed extensive networks. The undifferentiated cells did not exhibit a neuronal or glial phenotype. The differentiating cells expressed nestin, a marker of neural stem cells, at 6 h, and disappeared at 6 d. In contrast, expression of neuron-specific markers and gene was detectable at 6 d. And glia-specific markers and gene was not expressed. The percentage of apoptotic cells was 12.2% +/- 2.8%. In addition, fluorescently labeled cells were readily detected in the brain section after 14d of transplantation. CONCLUSION: Baicalin may induce adult rat MSCs to neuron in vitro. It is a useful vehicles for autotransplantation in both cell and gene therapy for a variety of diseases of the central nervous system.

Animals↗

PubMed2002

Three-dimensional quantitative structure-activity relationship analysis of ligand binding to human sequence antidigoxin monoclonal antibodies using comparative molecular field analysis.

The present study indicates that the newly generated human sequence antidigoxin monoclonal antibody (mAb), 1B3, binds digoxin with a different fine specificity binding than our previously obtained human sequence monoclonal antibodies (mAbs) (Ball, W. J.; et al. J. Immunol. 1999, 163, 2291-2298). Uniquely, 1B3 has a higher affinity for digitoxin than digoxin, the immunizing hapten, and a strong requirement for at least one sugar residue linked to the aglycone (-genin). By means of comparative molecular field analysis (CoMFA), the results of competition binding studies for 56 cardiotonic and hormonal steroids were employed to develop three-dimensional quantitative structure-activity relationship (3D-QSAR) models for ligand binding to 1B3 and to three additional human sequence mAbs, as well as the murine antidigoxin mAb 40-50 (Mudgett-Hunter, M.; et al. Mol. Immunol. 1985, 22, 447-488). All five 3D-QSAR models yielded cross-validated q(2) values greater than 0.5, which indicates that they have significant predictive ability. The CoMFA StDevCoeff contour plots, as well as the competition results, indicate that 1B3 binds ligands in a manner distinct from the other four mAbs. The CoMFA contour plots for 40-50 were also compared with the known X-ray crystallographic structure of the 40-50-ouabain complex (Jeffrey, P. D.; et al. J. Mol. Biol. 1995, 248, 344-360) in order to identify correlations between residues in the mAb binding site and specific contour plot regions. These 3D-QSAR models and their respective contour plots should be useful tools to further understand the molecular nature of antibody-antigen interactions and to aid in the redesign or enhancement of therapeutic antibodies.

Amino Acid Sequence↗

PubMed2002

Endo-oxacyclizations of polyepoxides: biomimetic synthesis of fused polycyclic ethers.

Boron trifluoride-etherate promotes the endo-selective oxacyclization of polyepoxides derived from various acyclic terpenoid polyalkenes, including geraniol, farnesol, and geranylgeraniol, providing an efficient and stereoselective synthesis of substituted oxepanes and fused polyoxepanes. The mechanism of the oxacyclization reaction probably involves intramolecular nucleophilic addition of epoxide oxygen to open another epoxide that is activated as an electrophile by the Lewis acid. These oxacyclizations proceed stereospecifically with inversion of configuration upon opening of each epoxide to provide trans-fused polycyclic products. The oxacyclization cascade is terminated by a tethered nucleophile, which may be the carbonyl oxygen of a ketone, ester, or carbonate, or a trisubstituted alkene. The best oxacyclization yields are generally observed with tert-butyl carbonate as the terminating nucleophile, although in some cases the oxacyclization products include formation of tert-butyl ethers as a minor product. The oxacyclization transformations described herein may mimic ring-forming steps in the biosynthesis of trans-syn-trans-fused polycyclic ether marine natural products.

Biological Factors↗

PubMed2002

Computational studies on tetrahydropyrimidine-2-one HIV-1 protease inhibitors: improving three-dimensional quantitative structure-activity relationship comparative molecular field analysis models by inclusion of calculated inhibitor- and receptor-based properties.

A computational chemistry study has been performed on a series of tetrahydropyrimidine-2-ones (THPs) as HIV-1 protease (HIV-1 PR) inhibitors. The present investigation focuses on the correlation of inhibitor-enzyme complexation energies (E(compl)), inhibitor solvation energies E(solv)[I], and both polar and nonpolar buried surface areas (BSAs) with the observed values of the binding affinity (pK(I)). Various combinations of these specific inhibitor- and receptor-based properties were also evaluated as additional descriptors to three-dimensional quantitative structure-activity relationship (3D-QSAR) models constructed using comparative molecular field analysis (CoMFA). Linear regression of the observed pK(I) values with E(compl), E(solv)[I], and the BSAs yielded a strong correlation in terms of both self-consistency (r(2) approximately equal to 0.90) and internal predictive ability (r(cv)(2) > 0.50). The 3D-QSAR models obtained from CoMFA using standard partial least-squares (PLS) analysis also yielded a strong correlation between the CoMFA fields and the experimental pK(i) (r(2) = 0.96; r(cv)(2) = 0.58). Various "enhanced" 3D-QSAR models were constructed in which different combinations of the E(compl), E(solv)[I], and BSAs were added as additional descriptors to the default steric-electrostatic CoMFA fields. Inclusion of E(solv)[I] in particular yielded significant improvement in the predictive ability (r(cv)(2) approximately equal to 0.80) of the resultant 3D-QSAR model.

HIV Protease↗

PubMed2002

Three-dimensional quantitative structure-activity relationship study of the inhibition of Na(+),K(+)-ATPase by cardiotonic steroids using comparative molecular field analysis.

Na(+),K(+)-ATPase is a transmembrane protein that transports sodium and potassium ions across cell membranes during an activity cycle that uses the energy released by ATP hydrolysis. Cardiotonic steroids (digitalis) inhibit this activity and consequently produce a positive inotropic response in the heart. To identify the structural features of the steroids that are important for this inhibition, we have tested the inhibitory properties of 47 cardiotonic and hormonal steroids and developed a three-dimensional quantitative structure-activity relationship (3D-QSAR) model for the inhibition of Na(+),K(+)-ATPase using comparative molecular field analysis (CoMFA). We also developed a 3D-QSAR model for the binding of digoxin to the murine anti-digoxin monoclonal antibody (mAb) 26-10 because we have previously shown that the environment of the binding sites of 26-10 and the enzyme are similar (Kasturi et al. (1998) Biochemistry 37, 6658-6666). These statistically predictive 3D-QSAR models indicate that both binding sites are about 20 A long and have a close fit or complementarity about the beta side of the lactone ring of digitalis. Furthermore, steric bulk about the lactone ring and the alpha sugar may be critical for drug binding. However, the binding site of Na(+),K(+)-ATPase differs from that of mAb in that it has a greater number of electrostatic interactions along the alpha-sugar, steroid, and lactone moieties. In addition, the availability of the structure of the 26-10 Fab-digoxin complex (Jeffrey et al. (1993) Proc. Natl. Acad. Sci. U.S.A. 90, 10310-10314) enabled us to compare the CoMFA-derived contour maps with the known locations for amino acid residues comprising the mAb ligand binding site.

Animals↗

PubMed2002

Vitamin C and vitamin E supplementation reduce oxidative stress-induced embryo toxicity and improve the blastocyst development rate.

To evaluate the adverse effects of exogenously induced reactive oxygen species (ROS) on mouse embryo development by using the 12-phorbol 13-myristate acetate (PMA)-activated leukocyte model as a source of ROS, and to examine the protective effect of antioxidant supplementation (vitamin C and vitamin E). Prospective study. Research laboratory. Effects of ROS on the blastocyst development rate in the presence and absence of antioxidant supplementation. After incubation with the PMA-activated leukocyte supernatant, the median (25th, 75th percentile) rate of blastocyst development significantly decreased from 73% (60%, 80%) after 3 hours to 30% (20%, 40%) after 6 hours compared with control reactions (86% [80%, 100%]). Co-incubating the embryos with vitamin C (50 microM) and the PMA-activated supernatant significantly increased the blastocyst development rate from 73% (60%, 80%) to 90% (80%, 91%) at 3 hours and from 30% (20%, 40%) to 91% (89%, 91%) at 6 hours-a level similar to that of control reactions. The blastocyst development rate increased after vitamin E supplementation (400 microM) at 6 hours, but not significantly and not by as much as after vitamin C supplementation. Exposure of mouse embryos to ROS for extended periods results in embryotoxicity. Vitamin C is more effective than vitamin E in reversing ROS-induced mouse embryo toxicity.

Animals↗

PubMed2002

Relationship between oxidative stress and embryotoxicity of hydrosalpingeal fluid.

BACKGROUND: Oxidative stress mechanisms are involved in the pathophysiology of many reproductive disorders. The objective of this study was to characterize oxidative stress parameters in hydrosalpingeal fluid (HSF) and examine their possible role in early embryo development. METHODS AND RESULTS: HSF was aspirated at laparoscopic salpingectomy in 11 infertile women. Reactive oxygen species (ROS), total (non-enzymatic) antioxidant capacity (TAC) and lipid peroxidation (LPO) were assayed. Two-cell mouse embryos were incubated with 25, 50 or 75% HSF and the blastocyst development rate was observed. ROS was detected in five of 11 (45%) HSF samples with a mean of 4.2 x 10(4) c.p.m. LPO was detected in all samples at a mean (+/- SD) value of 5575.4 +/- 6091.9 micromol/l malonaldehyde. The mean blastocyst development rate at 25, 5+/- 0 and 75% HSF and in the control group was 88.9 +/- 9.4, 65.7 +/- 19.1, 45.7 +/- 5.7 and 96.7% respectively (P < 0.0001). The blastocyst development rate was positively correlated to ROS concentrations (P < 0.02) but was not significantly related to LPO. CONCLUSIONS: The blastocyst development rate decreased with increasing concentrations of HSF. For the first time, the presence of ROS, LPO and TAC activity in human HSF was characterized. A possible role of oxidative stress in the embryotoxicity of HSF is suggested.

Animals↗

PubMed2002

Sodium channel-blocking agents are not of benefit to rats with kaolin-induced hydrocephalus.

OBJECTIVE: Hydrocephalus causes damage to periventricular white matter at least in part through chronic ischemia. The sodium channel-blocking agents mexiletine and riluzole have been shown to be of some protective value in various models of neurological injury. We hypothesized that these agents would ameliorate the effects of experimental childhood-onset hydrocephalus. METHODS: Hydrocephalus was induced in 4-week-old rats by injection of kaolin into the cisterna magna. Tests of cognitive and motor function were performed on a weekly basis. In a blinded and randomized manner, mexiletine (0.7, 7, or 42 mg/kg/d) or riluzole (1.4 or 13.6 mg/kg/d) was administered by osmotic minipump for 2 weeks, beginning 2 weeks after induction of hydrocephalus. The brains were then subjected to histopathological and biochemical analyses. RESULTS: Compared with untreated hydrocephalic rats, neither mexiletine nor riluzole was associated with a protective effect on behavioral, structural, or biochemical abnormalities. CONCLUSION: Protection of hydrocephalic brains through pharmacological sodium channel blockade is probably an approach not worth pursuing.

Animals↗

PubMed2002

[Immune responses of dendritic cells after loaded with cytotoxicity T lymphocyte epitope based peptide of human alpha-fetoprotein (hAFP)].

OBJECTIVE: To study the immune responses of lymphocytes after activated by dendritic cells (DCs) loaded with cytotoxicity T lymphocyte (CTL) epitope based peptide of human alpha-fetoprotein (hAFP, 218-226 LLNQHACAV). METHODS: Get high purity DCs by cultured plastic-adherent monocytes isolated from healthy donor of HLA-A2(+) peripheral blood with granulocyte-monocyte colony-stimulating factor (GM-CSF) and interleukin-4 (IL-4) for 7 days. Stimulate self-lymphocytes with DCs that loaded with CTL epitope based peptide of hAFP under the culture medium contains interleukin-2 (IL-2) for 7 days. Analyse IL-12 and TNF in culture medium and also the specific lysis activity of lymphocytes against four strains of primary hepatocellular carcinoma cells. RESULTS: After stimulated by DC loaded with CTL epitope based peptide derived from hAFP, lymphocytes appeared a good characteristics and the culture medium of activated lymphocytes contained a high level Th1 type cytokines of IL-12 and TNF. Activated lymphocytes not only specifically lysed HLA-A2(+) HepG2 line but also had the cytotoxicity against other three primary hepatocellular carcinoma cell lines and T2 target cell loaded with peptide of hAFP. CONCLUSIONS: The results of this research supply the basic materials for the DC based vaccine with HLA-A2 restricted peptide epitope derived from hAFP against AFP positive primary hepatocellular carcinoma.

Adult↗