We evaluated the ability of lactic acid bacteria, Weissella cibaria, isolated from the oral cavity to adhere to epithelial cells. W. cibaria efficiently adhered to KB cells and HeLa cells. In addition, W. cibaria efficiently adhered to Fusobacterium nucleatum. But the adhesiveness of W. cibaria disappeared upon exposure to LiCl or pronase, suggesting that the S-layer proteins of W. cibaria mediated the adhesiveness. The molecular mass of the S-layer proteins extracted from W. cibaria was approximately 50 kDa. When W. cibaria strains were washed with 0.45% saline, the bacteria were efficiently adhered to the epithelial cells. In conclusion, W. cibaria has the ability to adhere to epithelial cells through the S-layer proteins.
Adhesiveness ; Bacteria ; Epithelial Cells* ; Fusobacterium nucleatum ; HeLa Cells ; Humans ; KB Cells ; Lactic Acid ; Mouth ; Pronase ; Weissella*

Cis-dichlorodiammine platinum II (Cisplatin), an effective chemotherapeutic agent, induces acute renal failure by unknown mechanisms. To investigate direct toxic effects of cisplatin on the renal proximal tubular transport system, LLC-PK-1 cell line was selected as a cell model and the sugar transport activity was evaluated during a course of cisplatin treatment. Cells grown to confluence were treated with cisplatin for 60 min, washed, and then incubated for up to 5 days. At appropriate intervals, cells were tested for sugar transport activity using alpha-methyl-D-(14C)glucopyranoside (AMG) as a model substrate. In cells treated with 100 micrometer cisplatin, the AMG uptake was progressively impaired after 3 days. The viability of cells was not substantially changed with cisplatin of less than 100 micrometer, but it decreased markedly with 150 and 200 micrometer. In cisplatin-treated cells, the Na+/-dependent AMG uptake was drastically inhibited with no change in the Na+/-independent uptake. Kinetic analysis indicated that Vmax was suppressed, but Km was not altered. The Na+/-dependent phlorizin binding was also decreased in cisplatin-treated cells. However, the AMG efflux from preloaded cells was not apparently retarded by cisplatin treatment. These data indicate that the cisplatin treatment impairs Na+/-hexose cotransporters in LLC-PK-1 cells and suggest strongly that defects in transporter function at the luminal plasma membrane of the proximal tubular cells constitute an important pathogenic mechanism of cisplatin nephrotoxicity.
Acute Kidney Injury ; Animals ; Cell Line ; Cell Membrane ; Cisplatin* ; Epithelial Cells* ; LLC-PK1 Cells ; Phenobarbital ; Phlorhizin ; Platinum ; Swine

OBJECTIVE: The purpose of the study was to determine the effects of co-culture with oviductal epithelial cells and Vero cells on mouse embryo. METHOD: For the control group, mouse embryos were cultured alone in Ham's F-10 with 10% FBS. Subcultured oviductal epithelial cell and Vero cell were cocultured in Ham's F-10 with 10% FBS with the mouse embryo and used as the treatment group. Development of mouse embryos were observed. Result: The development rate and hatching rate of embryos that cocultured with oviductal epithelial cell and Vero cell was significantly higher (p<0.05) than control group. When subcultured oviductal epithelial cells were co-cultured with mouse embryo, there was no significant difference in development rate and hatching rate among subculture step. When oviductal epithelial cells that have been frozen-thawed were co-cultured with mouse embryo, there was no significant difference in development rate and hatching rate among subculture step. No statistical significance was seen in the development rate and hatching rate between subcultured oviductal epithelial cells and frozen-thawed oviductal epithelial cells when cocultured with mouse embryo, Vero cells and frozen-thawed when cocultured with mouse embryo, and Vero cells and oviductal epithelial cells when cocultured with mouse embryo. CONCLUSION: Oviductal epithelial cells and Vero cell may have a stimulatory role in early mouse embryonal development compared to control in vitro. As well, there is no significant difference in development rate and hatching rate among subculture step, when early mouse embryo was cocultured with cells that subcultured and frozen-thawed.
Animals ; Coculture Techniques ; Embryonic Structures ; Epithelial Cells* ; Humans* ; Mice* ; Oviducts* ; Vero Cells*

OBJECTIVETo investigate the way of herpes simplex virus type 1 (HSV-1) infecting human oral epithelial cell in vitro.

METHODSAbundance of HSV-1 strains were used to infect human oral epithelial cells. The culture supernatant was collected to infect Vero cells again. Morphology of HSV-1 was identified by inverted microscope and transmission electron microscope. Nucleic acid of the virus was detected by PCR.

RESULTSThe infected human oral epithelial cells didn't display obvious cytopathic effect (CPE) while Vero cells infected by the culture supernatant showed typical CPE. The virus particles were observed in Vero cells. The nucleic acid of HSV-1 could be detected in infected human oral epithelial cells by PCR.

CONCLUSIONSHSV-1 could successfully infect human oral epithelial cells.

Animals ; Cercopithecus aethiops ; Epithelial Cells ; virology ; Herpesvirus 1, Human ; pathogenicity ; Humans ; Mouth ; cytology ; Vero Cells

This study was purposed to investigate the relationship between the catalysis of Bence Jones protein (BJP) in urine of patients with multiple myeloma(MM) and toxicity on the renal proximal tubular cells in vitro, and to explore the potential mechanism for the toxicity of BJP to renal impairment in patients with MM. The Michaelis-Menten constant (K(m)) and catalytic constant (k(cat)) of the amidase activity of BJP was calculated by Hanes equation. The LLC-PK1 cells were cultured with different concentration of BJP for 24 h, then proliferation of the cells were determined by MTT method and apoptosis were determined by flow cytometry. The results showed that the BJP from the MM patients with renal impairment significantly inhibited cell proliferation, as compared with that from MM patients without renal impairment. The BJP with higher k(cat) had higher toxicity to LLC-PK1 cells. BJP could induce apoptosis and necrosis of LLC-PK1 cells when reached a certain concentration and this effect enhanced with increase of BJP concentration. It is concluded that the catalysis of BJP and its toxicity to renal tubular epithelial cells has a positive correlation, and toxic effect of BJP on renal tubular epithelial cells results from inhibiting proliferation and inducing apoptosis and necrosis of the cells, which may be one of renal impairment mechanisms in MM patients.
Animals ; Bence Jones Protein ; metabolism ; toxicity ; Catalysis ; Coculture Techniques ; Epithelial Cells ; metabolism ; pathology ; Humans ; Kidney ; metabolism ; pathology ; Kidney Tubules ; cytology ; LLC-PK1 Cells ; Multiple Myeloma ; metabolism ; pathology ; Swine

BACKGROUND: 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), a prototype compound of polyhalogenated aromatic hydrocarbons, produces diverse biologic effects. Although nephrotoxicity of aromatic hydrocarbons such as benzo[a]pyrene(BP) is well known, little is known about the effects of TCDD on renal function. Thus, the present study examined the effects of TCDD on cell viability, proliferation, and extracellular matrix(ECM) synthesis by glomerular mesangial cells, LLC-PK1 cells representing proximal tubular epithelial cells, and MDCK cells representing distal epithelial cells and compared with the effects of BP. METHODS: Quiescent cells were incubated with serum free media containing different concentrations of TCDD(1-100 nM) and BP(3 and 30 micro M) for 24- 96 hours. Cell viability and proliferation were assessed by lactate dehydrogenase(LDH) release and [3H]-thymidine incorporation, respectively. Secreted fibronectin was measured by Western blot analysis. RESULTS: When cells were continuously exposed to TCDD, LDH release significantly increased in MMC, LLC-PK1, and MDCK in a dose- and a time- dependent manner. [3H]-Thymidine incorporation was increased in MMC and LLC-PK1 but decreased in MDCK by TCDD. Contrary to TCDD, 30 micro BP significantly inhibited [3H]-thymidine incorporation in MMC and MDCK but not in LLC-PK1. Both TCDD and BP increased fibronectin secretion by MMC, LLC-PK1, and MDCK cells, suggesting that TCDD and BP may cause renal fibrosis leading to loss of renal function. CONCLUSION: These data provide experimental evidence that TCDD can alter cell viability and proliferation and increase ECM synthesis by renal cells which may lead to renal injury.
Animals ; Blotting, Western ; Cell Survival ; Culture Media, Serum-Free ; Epithelial Cells* ; Fibronectins ; Fibrosis ; Hydrocarbons, Aromatic ; Lactic Acid ; LLC-PK1 Cells ; Madin Darby Canine Kidney Cells ; Mesangial Cells ; Swine ; Tetrachlorodibenzodioxin

OBJECTIVETo detect the expression changes of the demethylase TETs (Ten-eleven translocation enzymes) in human embryonic kidney cell (HEK293) exposed to high dose cadmium chloride (CdCl2), and to investigate the regulation effects of TETs on global genomic methylation.

METHODSHEK293 cells were exposed to CdCl2 for 24 h, 48 h and 72 h, the survival rate was tested by CCK-8 (cell counting kit-8) method, and the cell morphology was observed. The levels of TETs mRNA and protein were detected by fluorescence quantitative PCR and Western blot, respectively. The genomic DNA methylation level was detectedby pyro sequencing assay.

RESULTSCdCl2 had toxic effects on HEK293 cells, and the half inhibitory concentration (IC50) was 1.78 µmol/L. After exposure of CdCl2 for 24 h, 48 h and 72 h, the morphology of HEK293 cells was altered, and the high dose group (2.0 µmol/L) showed vacuolar changes and fuzzy appearance. The level of TET1 mRNA in groups of 0.0, 0.5, 1.0, 2.0 µmol/L were 0.23 ± 0.13, 0.48 ± 0.12, 0.59 ± 0.16 and 0.95 ± 0.39, respectively (F = 182.89, P = 0.002); The level of TET2 mRNA in groups of 0.0, 0.5, 1.0, 2.0 µmol/L were 0.23 ± 0.12, 0.32 ± 0.02,0.31 ± 0.10 and 0.34 ± 0.07, respectively (F = 27.94, P < 0.001); The level of TET3 mRNA in groups of 0.0, 0.5, 1.0, 2.0 µmol/L were 0.26 ± 0.10, 0.27 ± 0.11, 0.25 ± 0.11 and 0.28 ± 0.09, respectively (F = 1.76, P = 0.036). The interaction effect existed between exposure time and doses of TET1 mRNA, TET2 mRNA and TET3 mRNA (F values were 32.94, 23.04 and 13.78, respectively; P values were < 0.001, 0.041 and < 0.001, respectively). Western blot showed that in different exposure time and dose, the protein expression levels of TETs had the similar trend as mRNA levels. In 24 h (55.01 ± 3.62)%, 48 h (48.31 ± 8.99)%, 72 h (48.76 ± 6.60)%, the DNA methylation had significant differences (F = 18.50, P < 0.001); In groups of 0.0 µmol/L (55.29 ± 2.83)%, 0.5 µmol/L (55.35 ± 3.11)%, 1.0 µmol/L (48.58 ± 6.40)% and 2.0 µmol/L (43.56 ± 7.89)%, the differences of DNA methylation had significant differences (F = 7.03, P = 0.048); the effect of interaction was also existed (F = 2.73, P = 0.043).

CONCLUSIONIn the short term exposure to CdCl2, the levels of TETs mRNA and protein showed a trend of increase according to the exposure time and dose, and the methylation level of whole genomic DNA was also altered. The demethylase TETs may play a role in regulating the genomic methylation level of HEK293 exposed to cadmium.

Cadmium Chloride ; toxicity ; DNA Methylation ; Dioxygenases ; genetics ; Epithelial Cells ; drug effects ; HEK293 Cells ; Humans ; RNA, Messenger

Persistent cell volume reduction is a major hallmark of apoptosis. Recent studies have demonstrated that cell volume reduction is not a passive, secondary event of the apoptotic cell death process. Whole-cell shrinkage, termed apoptotic volume decrease (AVD), takes place soon after stimulation with apoptogen and precedes caspase activation, DNA and cell fragmentation in a variety of cell types including human epithelial cells. The AVD induction is the result of KCl efflux attained by activation of K(+) and Cl(-) channels. Inhibition of AVD induction leads to rescue of the cells from apoptosis. Since the AVD process is coupled to dysfunction of the regulatory volume increase (RVI), apoptotic cells undergo persistent cell shrinkage in human epithelial HeLa cells. When the RVI mechanism was impaired, hypertonic stress itself induced not only persistent cell shrinkage but also apoptotic cell death in HeLa cells. Even under normotonic apoptogen-free conditions, exposure of HeLa cells to Na(+)- or Cl(-)-deficient solution alone can bring about persistent cell shrinkage and thereafter apoptotic cell death. Thus, it is concluded that persistent cell shrinkage, which comprises AVD induction and RVI dysfunction, is a prerequisite to apoptosis induction in human epithelial cells.
Apoptosis ; Cell Size ; Epithelial Cells ; cytology ; HeLa Cells ; Humans ; Osmotic Pressure

To investigate the inflammatory cytokine production of human epithelial cell lines stimulated with uropathogenic E. coli strains showing 3 different adherence patterns, differential expression of inflammatory cytokine (IL-1a, IL-lB, IL-8, TNFa, and TGFB) mRNA were detected by RT-PCR. IL-1a, IL-1B, IL-8, and TGFB mRNAs constitutively expressed in epithelial cell lines, but not TNFa. The expression of IL-1a and IL-1B mRNA was increased in J-82 cells stimulated with E. coli strains showing DA, LA, or AggA pattern. The expression of IL-8 mRNA was increased, whereas TGFj3 mRNA was decreased in J-82 cells stimulated with E. coli strain showing AggA.pattern. Treatment with crude bacterial adhesins (CBA) isolated from E. coli strains showing DA or LA pattern increased IL-la, IL-lB, IL-S, and TGFj3 mRNA expressions in J-82 cells and HeLa cells. IL-la, IL-lB, and TGFB mRNA expressions were decreased in epitheUal cells stimulated with CBA from E. coli strain showing AggA pattern, whereas IL-8 mRNA expression was significantly increased. The expressions of cytokine mRNAs showed little differences between epithelial ceRs used, but great differences between CBA from DA or LA and AggA strain. LPS stimulation was little changed cytokine mRNA expressions in epithelial cells. This study suggests that cytokine gene expression of epithelial cells by the bacterial stimulation mainly depends on the bacterial adhesins recognized by the respective receptors of epithelial cells.
Adhesins, Bacterial ; Epithelial Cells* ; Gene Expression ; HeLa Cells ; Humans ; Interleukin-8 ; RNA, Messenger ; Uropathogenic Escherichia coli*

Brucellosis is one of the common zoonoses caused by Brucella abortus (B. abortus). However, little has been reported on factors affecting invasion of B. abortus into host cells. To investigate cell-type dependent invasion of B. abortus, phagocytic RAW 264.7 and THP-1 cells and non-phagocytic HeLa cells were infected with wild-type and mutant B. abortus, and their invasion efficiencies were compared. The invasion efficiencies of the strains were cell-type dependent. Wild-type B. abortus invasion efficiency was greater in phagocytic cells than in epithelial cells. The results also indicated that there are different factors involved in the invasion of B. abortus into phagocytic cells.
Brucella abortus ; Brucella ; Brucellosis ; Epithelial Cells ; HeLa Cells ; Humans ; Phagocytes ; Zoonoses