Bacterial attachment to oropharyngeal epithelial cells in breastfed newborns

Dopaminergic agents and carbidopa/levodopa have become the preferred treatment for both the restless legs (RL) syndrome and for periodic limb movements in sleep (PLMS). For once-nightly treatments with carbidopa/ levodopa, a problem with morning end-of-dose rebound increases in leg movements has been reported to occur in the about one-fourth of the patients. In our clinical studies a previously unreported but far more significant problem of markedly augmented RL symptoms occurred in the afternoon and the evening prior to taking the next nightly dose. A systematic prospective evaluation of this augmentation in 46 consecutive patients treated with carbidopa/ levodopa for RL syndrome or PLMS disorder found this augmentation to be the major adverse effect of treatment. Augmentation occurred for 31% of PLMS patients and 82% of all RL patients. It was greater for subjects with more severe RL symptoms and for patients on higher doses (> or = 50/200 mg carbidopa/levodopa) but was unrelated to gender, age or baseline severity of PLMS. This augmentation was severe enough to require medication change for 50% of the RL patients and 13% of PLMS patients. Augmentation resolved with cessation of the medication and could be minimized by keeping the dose low.     

Proliferation of corneal epithelial cells in diabetic rats

In order to elucidate the mechanisms of diabetic corneal epitheliopathy, we investigated the proliferation of corneal epithelial cells of streptozotocin-induced diabetic rats in the 1st, 2nd, 3rd and 6th months after the onset of diabetes using 3H-thymidine autoradiography. After the 1st month in diabetic rats, histological examination revealed that corneal epithelial layer was thin, and proliferation of corneal epithelial cells was decreased compared with that of normal untreated rats. After the 2nd month, proliferation of corneal epithelial cells showed no difference from normal rats. After the 3rd month in diabetic rats, the attachment of the epithelial layer to the stroma was loose and proliferation of corneal epithelial cells was increased compared with that of normal untreated rats. After the 6th month, proliferation of corneal epithelial cells showed no difference between diabetic rats and normal rats. We suggest that increased turnover rate of corneal epithelial cells may account for increased proliferation of corneal epithelial cells in diabetic rats during the 3rd month.     

Multiple attachment mechanisms of corneal epithelial cells to a polymer--cells can attach in the absence of exogenous adhesion proteins through a mechanism that requires microtubules

STUDY OBJECTIVE: To determine whether emergency patients with acute chest pain and low suspicion of acute myocardial infarction (AMI) can be managed cost-effectively and safely in a dedicated chest pain center (CPC) that incorporates mandatory stress testing. METHODS: We assembled a prospective observational case series of consecutive adult patients transferred from the emergency department to a nine-bed, 23-hour CPC in a 564-bed community hospital from January 13 through May 31, 1994. In our institution, all emergency patients with acute nontraumatic chest pain of unclear origin, suggestive of myocardial ischemia but with a low probability of AMI, are transferred to the CPC for further evaluation. All patients in whom AMI is ruled out undergo individually appropriate cardiac diagnostic testing in accordance with CPC clinical guidelines. Patients with end-stage coronary artery disease transferred to the CPC for a "rule-out" protocol only did not undergo further diagnostic testing. Admitted and discharged patients were followed through chart review and telephone survey, respectively. RESULTS: Of the 502 patients transferred to the CPC, 477 (95%) completed follow-up at 14 days. Four hundred ten (86%) were discharged home. Those discharged after diagnostic evaluation yielded negative findings had 100% survival and zero diagnosis of AMI at 5-month follow-up. Overall mortality and incidence of AMI on long-term follow-up for all patients transferred to the CPC were .4% and .2%, respectively. Sixty-seven patients (13%) were admitted from the CPC, of whom 44 (66%) had a final diagnosis of ischemic heart disease (IHD) or AMI. Twenty-four patients with IHD (55%; 6% of stress-tested group) were identified only on further stress testing. Of these patients, seven underwent percutaneous transluminal coronary angioplasty or coronary artery bypass grafting during hospitalization. All were discharged home without major morbidity. Four hundred twenty-four patients (84%) underwent stress testing. The cost of mandatory stress testing to identify one patient with IHD after AMI was ruled out was $3,125. An average cost-per-case savings of 62% was achieved for each patient transferred to the CPC who would have been hospitalized before the inception of the CPC. CONCLUSION: Mandatory stress testing is a safe, cost-effective, and valuable diagnostic and prognostic tool in CPC patients.     

Rabbit corneal epithelial cells grown in vitro without serum

Primary outgrowth cultures of normal rabbit corneal epithelium can be initiated and propagated in vitro up to 6 days in serum-free medium. By the eighth day the majority of cells have ceased to divide. Epithelial cells grown without serum show DNA synthetic activity at a level comparable to control cultures grown with added serum.     

Roxithromycin inhibits cytokine production by and neutrophil attachment to human bronchial epithelial cells in vitro

We evaluated the effect of roxithromycin on cytokine production and neutrophil attachment to human airway epithelial cells. Roxithromycin suppressed production of interleukin 8 (IL-8), IL-6, and granulocyte-macrophage colony-stimulating factor. It inhibited neutrophil adhesion to epithelial cells. Roxithromycin modulates local recruitment and activation of inflammatory cells, which may have relevance to its efficacy in airway diseases.     

Attachment of Burkholderia pseudomallei to pharyngeal epithelial cells: a highly pathogenic bacteria with low attachment ability

OBJECTIVE: To identify the mutation in a human transforming growth factor beta-induced gene (BIGH3) in a Japanese family with a severe form of granular corneal dystrophy of early onset associated with recurrent corneal erosions. PATIENTS: The tentative clinical diagnosis in this family was Reis-Bücklers corneal dystrophy; 4 persons affected with this disorder have been identified in 4 generations, and 3 of the 4 were examined. The proband underwent keratoplasties in our hospital (Keio University Hospital, Tokyo, Japan). METHODS: The BIGH3 gene was examined for a mutation by the polymerase chain reaction and direct sequencing. Corneal buttons of the proband were stained and examined by electron microscopy. RESULTS: Three affected persons were shown to have a heterozygous G-->T transversion at the second nucleotide position of codon 124 (Arg-->Leu) of the BIGH3 gene. In the proband, corneal deposits between the epithelium and the Bowman layer stained red with Masson trichrome stain. Electron microscopy revealed numerous electron-dense, rod-shaped bodies next to the epithelial basement membrane but no curly fibers suggestive of Thiel-Behnke dystrophy. CONCLUSION: A novel R124L mutation of the BIGH3 gene was associated in this family with a superficial variant of granular corneal dystrophy. CLINICAL RELEVANCE: This mutation causes a severe form of superficial granular corneal dystrophy by producing abnormal keratoepithelin between the epithelium and the Bowman layer and thus clinical similarities to Reis-Bücklers corneal dystrophy.     

Ex vivo transduction of corneal epithelial progenitor cells using a retroviral vector

An in vivo model was used to determine whether bone hyperemia precedes increased intracortical porosity induced by disuse. Twenty-four adult male roosters (age 1 yr) were randomly assigned to intact-control, 7-days-sham-surgery, 7-days-disuse, and 14-days-disuse groups. Disuse was achieved by isolating the left ulna diaphysis from physical loading via parallel metaphyseal osteotomies. The right ulna served as an intact contralateral control. Colored microspheres were used to assess middiaphyseal bone blood flow. Bone blood flow was symmetric between the left and right ulnae of the intact-control and sham-surgery groups. After 7 days of disuse, median (+/-95% confidence interval) standardized blood flow was significantly elevated compared with the contralateral bone (6.5 +/- 5.2 vs. 1.0 +/- 0.8 ml x min-1 x 100 g-1; P = 0.03). After 14 days of disuse, blood flow was also elevated but to a lesser extent. Intracortical porosity in the sham-surgery and 7-days-disuse bones was not elevated compared with intact-control bones. At 14 days of disuse, the area of intracortical porosity was significantly elevated compared with intact control bones (0.015 +/- 0.02 vs. 0. 002 +/- 0.002 mm2; P = 0.03). We conclude that disuse induces bone hyperemia before an increase in intracortical porosity. The potential interaction between bone vasoregulation and bone cell dynamics remains to be studied.     

Calcium oxalate crystal attachment to cultured kidney epithelial cell lines

Lamprey spinal neurons exhibit a fast afterhyperpolarization and a late afterhyperpolarization (AHP) which is due to the activation of apamin-sensitive SK Ca2+-dependent K+ channels (KCa) activated by calcium influx through voltage-dependent channels during the action potential (Hill et al. 1992, Neuroreport, 3, 943-945). In this study we have investigated which calcium channel subtypes are responsible for the activation of the KCa channels underlying the AHP. The effects of applying specific calcium channel blockers and agonists were analysed with regard to their effects on the AHP. Blockade of N-type calcium channels by omega-conotoxin GVIA resulted in a significant decrease in the amplitude of the AHP by 76.2+/-14.9% (mean +/- SD). Application of the P/Q-type calcium channel blocker omega-agatoxin IVA reduced the amplitude of the AHP by 20.3+/-10.4%. The amplitude of the AHP was unchanged during application of the L-type calcium channel antagonist nimodipine or the agonist (+/-)-BAY K 8644, as was the compound afterhyperpolarization after a train of 10 spikes at 100 Hz. The effects of calcium channel blockers were also tested on the spike frequency adaptation during a train of action potentials induced by a 100-200 ms depolarizing pulse. The N- and P/Q-type calcium channel antagonists decreased the spike frequency adaptation, whereas blockade of L-type channels had no effect. Thus in lamprey spinal cord motor- and interneurons, apamin-sensitive KCa channels underlying the AHP are activated primarily by calcium entering through N-type channels, and to a lesser extent through P/Q-type channels.     

Substratum-dependent and region-specific control of attachment and proliferation of gastrointestinal epithelial cells in primary serum-free culture

A system for the primary serum-free culture of fetal rat gastrointestinal epithelial cells was used to examine the role of the extracellular matrix (ECM) in the attachment and proliferation of these epithelial cells. Forestomach epithelial cells (FSEC) were able to attach to and proliferate on plastic dishes without a substratum, while glandular stomach epithelial cells (GSEC) and duodenal epithelial cells (DEC) were unable to do so. The presence of a substratum promoted the attachment and proliferation of these epithelial cells. The effects of various components of the ECM differed depending on the type of cell. FSEC attached most efficiently to a substratum of fibronectin, while GSEC did so to laminin. DEC attached more efficiently to type I collagen and fibronectin than to any other substratum. FSEC proliferated most rapidly on laminin, while GSEC and DEC did so on collagen gels. These substrata induced the most efficient attachment and proliferation of FSEC, and they were effective in promoting the attachment and proliferation of GSEC and DEC in decreasing order of efficiency, indicating the existence of a head-to-tail gradient in the response of epithelial cells to substrata. The expression of c-myc mRNA in these cells differed depending upon the substratum on which they were cultured and the mRNA level was well correlated with the extent of the cell proliferation, indicating that the cell proliferation is mediated by c-myc gene expression, which is regulated by cell-ECM interactions. The results of the present study demonstrate that proliferation of gastrointestinal epithelial cells is regulated region-specifically not only by soluble factors but also by insoluble components of the ECM.     

Expression of integrin receptors on plasma membranes of primary corneal epithelial cells is matrix specific

The performance evaluation of a semi-supervised fuzzy c-means (SFCM) clustering method for monitoring brain tumor volume changes during the course of routine clinical radiation-therapeutic and chemo-therapeutic regimens is presented. The tumor volume determined using the SFCM method was compared with the volume estimates obtained using three other methods: (a) a k nearest neighbor (kNN) classifier, b) a grey level thresholding and seed growing (ISG-SG) method and c) a manual pixel labeling (GT) method for ground truth estimation. The SFCM and kNN methods are applied to the multispectral, contrast enhanced T1, proton density, and T2 weighted, magnetic resonance images (MRI) whereas the ISG-SG and GT methods are applied only to the contrast enhanced T1 weighted image. Estimations of tumor volume were made on eight patient cases with follow-up MRI scans performed over a 32 week interval during treatment. The tumor cases studied include one meningioma, two brain metastases and five gliomas. Comparisons with manually labeled ground truth estimations showed that there is a limited agreement between the segmentation methods for absolute tumor volume measurements when using images of patients after treatment. The average intraobserver reproducibility for the SFCM, kNN and ISG-SG methods was found to be 5.8%, 6.6% and 8.9%, respectively. The average of the interobserver reproducibility of these methods was found to be 5.5%, 6.5% and 11.4%, respectively. For the measurement of relative change of tumor volume as required for the response assessment, the multi-spectral methods kNN and SFCM are therefore preferred over the seedgrowing method.     

The effect of sodium hyaluronate on the growth of rabbit corneal epithelial cells in vitro

Rat spermatidal protein TP2 is a basic nuclear protein containing two atoms of zinc bound per molecule. We report here cloning of complementary DNA encoding rat TP2 by the RT-PCR method. The nucleotide sequence of cloned TP2 cDNA differs at a few positions from the sequence already reported in the literature. We have cloned rat TP2 cDNA into the expression vector pTrc 99A. Upon induction with 1 mM IPTG, there was a low level of expression of TP2 which could be recovered in the soluble form. Recombinant TP2 was purified from the soluble form. Recombinant TP2 was purified from the soluble extract of E. coli using nickel-agarose and heparin-agarose chromatography and was shown to be identical to native rat TP2 as revealed by immunoblotting with anti-rat TP2 antibodies and radioactive 65Zn-blotting.     

Effects of extracellular matrix constituents on the attachment of human oral epithelial cells at the titanium surface

This in vitro study attempts to delineate the role of extracellular matrix (ECM) constituents at the epithelial tissue-implant interface. To know which ECM constituents have a beneficial influence on the behavior of epithelial cells, the attachment, proliferation, morphologic pattern, and differentiation or cytoskeletal organization of human oral epithelial cells on ECM-coated (type IV collagen, fibronectin, type I collagen, laminin, and vitronectin) and noncoated titanium surface have been evaluated and compared. In each experiment comparing commercially pure titanium and oxygen plasma-cleaned titanium, the same ECM constituents were used. In this study, type IV collagen could provide an excellent substratum for epithelial cell attachment on titanium surface, but vitronectin-coated titanium revealed lower effectiveness for attachment of epithelial cells than noncoated titanium. These results suggested that type IV collagen could be used as a means for obtaining good epithelial seal, whereas vitronectin could be used to restrain the attachment of epithelium to dental implants.     

A murine and a porcine coronavirus are released from opposite surfaces of the same epithelial cells

Epithelial cells are important target cells for coronavirus infection. Earlier we have shown that transmissible gastroenteritis coronavirus (TGEV) and mouse hepatitis coronavirus (MHV) are released from different sides of porcine and murine epithelial cells, respectively. To study the release of these viruses from the same cells, we constructed a porcine LLC-PK1 cell line stably expressing the recombinant MHV receptor cDNA (LMR cells). The MHV and TGEV receptor glycoproteins were shown by immunofluorescence to appear at the surface of the cells and to be functional so that the cells were susceptible to both MHV and TGEV infection. Both coronaviruses entered polarized LMR cells only through the apical surface. Remarkably, while the cells remained susceptible to TGEV for long periods, infectability by MHV decreased with time after plating of the cells onto filters. This was not due to a lack of expression of the MHV receptor, since this glycoprotein was still abundant on the apical surface of these cells. TGEV and MHV appeared to exit LMR cells from opposite sides. Whereas TGEV was released preferentially at the apical membrane, MHV was released preferentially at the basolateral surface. These results show that vesicles containing the two coronaviruses are targeted differently in LMR cells. We propose that the viruses are sorted at the Golgi complex into different transport vesicles that carry information directing them to one of the two surface domains. The apical release of TGEV and the basolateral release of MHV might be factors contributing to the difference in virus spread found between TGEV and MHV in their respective natural hosts, the former causing mainly a localized enteric infection, the latter spreading through the body to other organs.     

Decreased corneal sensation as an initial feature of Acanthamoeba keratitis

BACKGROUND: Herpes simplex keratitis is the most common misdiagnosis in patients with Acanthamoeba keratitis, which is increasing in frequency and is associated with daily wear soft contact lenses. Both entities usually present as unilateral keratitis. The manifestations of superficial Acanthamoeba keratitis (i.e., unilaterality, dendriform appearance, positive response to antivirals, and decreased corneal sensation) increase the opportunity for misdiagnosis as herpes simplex keratitis. The authors have encountered six patients with Acanthamoeba keratitis in whom the correct diagnosis was delayed from 2 weeks to 3 months. METHODS: All six patients underwent testing with the Cochet-Bonnet esthesiometer and extensive pharmacologic treatment for herpes simplex keratitis. Corneal scrapings were taken between 2 and 6 weeks after the initial examination. RESULTS: In all six patients, corneal sensation was decreased significantly. Drug therapy was ineffective. Cultures were positive for Acanthamoeba. Five of six patients underwent penetrating keratoplasty. CONCLUSIONS: Decreased corneal sensation has contributed to the misdiagnosis of Acanthamoeba as herpes simplex keratitis. Misdiagnosis results in delayed treatment and worse outcome. The authors found that significantly decreased corneal sensation is a frequent finding in early Acanthamoeba keratitis. Therefore, physicians should consider Acanthamoeba keratitis as an alternative diagnosis in patients with presumed herpes simplex keratitis with decreased corneal sensation.     

Note: lack of influence of adherent Lactobacillus isolates on the attachment of Escherichia coli to the intestinal epithelial cells of chicken in vitro

Two Lactobacillus isolates, Lact. acidophilus I 26 and Lact. fermentum I 25, were selected, based on their poor aggregation with Escherichia coli and strong ability to adhere to ileal epithelial cells (IEC), to study in vitro interactions with E. coli O1:K1, O2:K1 and O78:K80 in an IEC radioactive-assay under the conditions of exclusion (lactobacilli and IEC, followed by the addition of E. coli), competition (lactobacilli, IEC and E. coli together) and displacement (E. coli and IEC, followed by the addition of lactobacilli). The results indicated that Lact. acidophilus I 26 and Lact. fermentum I 25 could not significantly reduce the attachment of E. coli O1:K1, O2:K1 and O78:K80 to IEC under the three conditions tested in vitro, except that the attachment of E. coli O1:K1 was slightly reduced by Lact. fermentum I 25 in the test for competition.     

Involvement of integrins with adhesion-promoting, heparin-binding peptides of type IV collagen in cultured human corneal epithelial cells

PURPOSE: The aim of this work was to identify the integrin subunits present on the cell surface of human corneal epithelial cells. The authors determined to show whether type IV collagen, heparin-binding peptides of type IV collagen (Hep-I, Hep-II, and Hep-III), fibronectin, and GRGDSP promote cell adhesion of human corneal epithelial cells. Type IV collagen and heparin-binding peptides of type IV collagen may be important in corneal epithelial cell adhesion in normal and pathologic conditions and reepithelialization. The authors assess the role of cell surface integrins in mediating cell adhesion to these proteins and peptides. METHODS: Fluorescence-activated cell sorter (FACS) analysis was used to determine the integrin subunits expressed at the cell surface of the cultured human corneal epithelial cells. Cell adhesion was assessed with type IV collagen, heparin-binding peptides of type IV collagen, fibronectin, and GRGDSP: Antibodies to the integrin subunits were used to determine the potential role of integrins in cell adhesion to the above proteins and peptides. RESULTS: FACS analysis identified the beta 1, beta 4, alpha 2, alpha 3, alpha 5, alpha 6, and alpha v integrin subunits on human corneal epithelial cells grown as primary cultures. The anti-beta 1 antibody inhibited cell adhesion to heparin-binding peptides of type IV collagen, type IV collagen, fibronectin, and GRGDSP: Antibodies to the alpha 2 integrin subunit inhibited cell adhesion to the heparin-binding peptides of type IV collagen and slightly inhibited cell adhesion to intact type IV. Antibodies to the alpha 3 integrin subunit exhibited a somewhat lesser effect compared to the anti-alpha 2 integrin antibody. CONCLUSIONS: These data show that the alpha 2 beta 1 integrin of human corneal epithelial cells recognize heparin-binding peptide sequences derived from human type IV collagen. It seems likely that these sequences play an important role in integrin-mediated corneal epithelial cell adhesion. In addition, the alpha 3 beta 1 integrin may mediate similar events.