A case of granulosa cell tumor of the ovary detected from metastatic foci.

The authors report a case of granulosa cell tumor of the ovary that followed a rare clinical course, where the primary focus did not appear as a mass, and disseminated foci grew in the abdominal cavity. In 2008, a 70-year-old patient, gravida 6 and para 3, was diagnosed with a perihepatic mass, peritoneal dissemination, and an abdominal wall mass as confirmed by computed tomography (CT) scanning. There was no mass lesion in the pelvis. The pathological diagnosis based on the resected mass in the abdominal wall was malignant mesothelioma. During follow-up, abdominal bloating developed from April 2009. CT scans indicated growth of the intraperitoneal lesions. Therefore, the patient received two cycles of combination therapy with cisplatin and pemetrexed. The treatment was discontinued due to lack of efficacy. The intraperitoneal lesions grew but the clinical course was slow and inconsistent with that of malignant mesothelioma. Central pathological review was requested in April 2011, and a granulosa cell tumor was diagnosed. The patient was referred to the department for detailed examination and treatment. The patient underwent incision of the intraperitoneal tumors, simple total hysterectomy, bilateral salpingo-oophorectomy, and omentectomy. The final pathological diagnosis was normal-size adult-type granulosa cell tumor originating from the left ovary. It was a case of granulosa cell tumor without ovarian enlargement where growth of the metastatic foci was the major observation. As complete surgical resection was achieved and no additional therapy was given, the subject was followed on an outpatient basis and no recurrence was identified.

Hepatitis A virus (HAV) proteinase 3C inhibits HAV IRES-dependent translation and cleaves the polypyrimidine tract-binding protein.

Hepatitis A virus (HAV) infection is still an important issue worldwide. A distinct set of viruses encode proteins that enhance viral cap-independent translation initiation driven by an internal ribosome entry site (IRES) and suppress cap-dependent host translation. Unlike cytolytic picornaviruses, replication of HAV does not cause host cell shut off, and it has been questioned whether HAV proteins interfere with its own and/or host translation. HAV proteins were coexpressed in Huh-7 cells with reporter genes whose translation was initiated by either cap-dependent or cap-independent mechanisms. Among the proteins tested, HAV proteinase 3C suppressed viral IRES-dependent translation. Furthermore, 3C cleaved the polypyrimidine tract-binding protein (PTB) whose interaction with the HAV IRES had been demonstrated previously. The combined results suggest that 3C-mediated cleavage of PTB might be involved in down-regulation of viral translation to give way to subsequent viral genome replication.

p19Arf sensitizes B16 melanoma cells to interferon-ß delivered via mesenchymal stem cells in vitro.

The immune stimulatory and anti-neoplastic functions of type I interferon have long been applied for the treatment of melanoma. However, the systemic application of high levels of this recombinant protein is often met with toxicity. An approach that provides localized, yet transient, production of type I interferon may overcome this limitation. We propose that the use of mesenchymal stem cells (MSCs) as delivery vehicles for the production of interferon-ß (IFNß) may be beneficial when applied together with our cancer gene therapy approach. In our previous studies, we have shown that adenovirus-mediated gene therapy with IFNß was especially effective in combination with p19Arf gene transfer, resulting in immunogenic cell death. Here we showed that MSCs derived from mouse adipose tissue were susceptible to transduction with adenovirus, expressed the transgene reliably, and yet were not especially sensitive to IFNß production. MSCs used to produce IFNß inhibited B16 mouse melanoma cells in a co-culture assay. Moreover, the presence of p19Arf in the B16 cells sensitizes them to the IFNß produced by the MSCs. These data represent a critical demonstration of the use of MSCs as carriers of adenovirus encoding IFNß and applied as an anti-cancer strategy in combination with p19Arf gene therapy.

The major laminin receptor of mouse embryonic stem cells is a novel isoform of the alpha 6 beta 1 integrin.

Laminin is the first extracellular matrix protein expressed in the developing mouse embryo. It is known to influence morphogenesis and affect cell migration and polarization. Several laminin receptors are included in the integrin family of extracellular matrix receptors. Ligand binding by integrin heterodimers results in signal transduction events controlling cell motility. We report that the major laminin receptor on murine embryonic stem (ES) cells is the integrin heterodimer alpha 6 beta 1, an important receptor for laminin in neurons, lymphocytes, macrophages, fibroblasts, platelets and other cell types. However, the cytoplasmic domain of the ES cell alpha 6 (alpha 6 B) differs totally from the reported cytoplasmic domain amino acid sequence of alpha 6 (alpha 6 A). Comparisons of alpha 6 cDNAs from ES cells and other cells suggest that the alpha 6 A and alpha 6 B cytoplasmic domains derive from alternative mRNA splicing. Anti-peptide antibodies to alpha 6 A are unreactive with ES cells, but react with mouse melanoma cells and embryonic fibroblasts. When ES cells are cultured under conditions that permit their differentiation, they become positive for alpha 6 A, concurrent with the morphologic appearance of differentiated cell types. Thus, expression of the alpha 6 B beta 1 laminin receptor may be favored in undifferentiated, totipotent cells, while the expression of alpha 6 A beta 1 receptor occurs in committed lineages. While the functions of integrin alpha chain cytoplasmic domains are not understood, it is possible that they contribute to transferring signals to the cell interior, e.g., by delivering cytoskeleton organizing signals in response to integrin engagement with extracellular matrix ligands. It is therefore reasonable to propose that the cellular responses to laminin may vary, according to what alpha subunit isoform (alpha 6 A or alpha 6 B) is expressed as part of the alpha 6 beta 1 laminin receptor. The switch from alpha 6 B to alpha 6 A, if confirmed in early embryos, could then be of striking potential relevance to the developmental role of laminin.

Laminin receptors in the retina: sequence analysis of the chick integrin alpha 6 subunit. Evidence for transcriptional and posttranslational regulation.

The integrin alpha 6 beta 1 is a prominent laminin receptor used by many cell types. In the present work, we isolate clones and determine the primary sequence of the chick integrin alpha 6 subunit. We show that alpha 6 beta 1 is a prominent integrin expressed by cells in the developing chick retina. Between embryonic days 6 and 12, both retinal ganglion cells and other retinal neurons lose selected integrin functions, including the ability to attach and extend neurites on laminin. In retinal ganglion cells, we show that this is correlated with a dramatic decrease in alpha 6 mRNA and protein, suggesting that changes in gene expression account for the developmental regulation of the interactions of these neurons with laminin. In other retinal neurons the expression of alpha 6 mRNA and protein remains high while function is lost, suggesting that the function of the alpha 6 beta 1 heterodimer in these cells is regulated by posttranslational mechanisms.

Epithelial integrin alpha 6 beta 4: complete primary structure of alpha 6 and variant forms of beta 4.

The integrin alpha 6 beta 4 is a heterodimer predominantly expressed by epithelia. While no definite receptor function has yet been assigned to it, this integrin may mediate adhesive and/or migratory functions of epithelial cells. We have determined the complete primary structure of both the alpha 6 and beta 4 subunits from cDNA clones isolated from pancreatic carcinoma cell line libraries. The deduced amino acid sequence of alpha 6 is homologous to other integrin alpha chains (18-26% identity). Antibodies to an alpha 6 carboxy terminus peptide immunoprecipitated alpha 6 beta 4 complexes from carcinoma cells and alpha 6 beta 1 complexes from platelets, providing further evidence for the association of alpha 6 with more than one beta subunit. The deduced amino acid sequence of beta 4 predicts an extracellular portion homologous to other integrin beta chains, and a unique cytoplasmic domain comprised of greater than 1,000 residues. This agrees with the structures of the beta 4 cDNAs from normal epithelial cells (Suzuki, S., and Y. Naitoh. 1990. EMBO [Eur. Mol. Biol. Organ.] J. 9:757-763; Hogervost, F., I. Kuikman, A. E. G. Kr. von dem Borne, and A. Sonnenberg. 1990. EMBO [Eur. Mol. Biol. Organ.] J. 9:765-770). Compared to these structures, however, the beta 4 cDNAs that we have cloned from carcinoma cells contain extra sequences. One of these is located in the 5'-untranslated region, and may encode regulatory sequences. Another specifies a segment of 70 amino acids in the cytoplasmic tail. Amplification by reverse transcription-polymerase chain reaction of mRNA indicated that multiple forms of beta 4 may exist, possibly due to cell-type specific alternative splicing. The unique structure of beta 4 suggests its involvement in novel cytoskeletal interactions. Consistent with this possibility, alpha 6 beta 4 is mostly concentrated on the basal surface of epithelial cells, but does not colocalize with components of adhesion plaques.

Integrin cytoplasmic domains mediate inside-out signal transduction.

We analyzed the binding of fibronectin to integrin alpha 5 beta 1 in various cells; in some cells fibronectin bound with low affinity (e.g., K562 cells) whereas in others (e.g., CHO), it bound with high affinity (Kd approximately 100 nM) in an energy-dependent manner. We constructed chimeras of the extracellular and transmembrane domains of alpha IIb beta 3 joined to the cytoplasmic domains of alpha 5 beta 1. The affinity state of these chimeras was assessed by binding of fibrinogen or the monoclonal antibody, PAC1. The cytoplasmic domains of alpha 5 beta 1 conferred an energy-dependent high affinity state on alpha IIb beta 3 in CHO but not K562 cells. Three additional alpha cytoplasmic domains (alpha 2, alpha 6A, alpha 6B) conferred PAC1 binding in CHO cells, while three others (alpha M, alpha L, alpha v) did not. In the high affinity alpha chimeras, cotransfection with a truncated (beta 3 delta 724) or mutated (beta 3(S752-->P)) beta 3 subunit abolished high affinity binding. Thus, both cytoplasmic domains are required for energy-dependent, cell type-specific affinity modulation. In addition, mutations that disrupted a highly conserved alpha subunit GFFKR motif, resulted in high affinity binding of ligands to alpha IIb beta 3. In contrast to the chimeras, the high affinity state of these mutants was independent of cellular metabolism, cell type, and the bulk of the beta subunit cytoplasmic domain. Thus, integrin cytoplasmic domains mediate inside-out signaling. Furthermore, the highly conserved GFFKR motif of the alpha subunit cytoplasmic domain maintains the default low affinity state.

Rationally Designed Peptide Probes for Extracellular Vesicles.

Extracellular vesicles (EVs) are submicroscopic lipid vesicles secreted from cells and play significant roles in cell-to-cell communication by transporting varieties of cell signaling molecules like proteins, DNA, mRNA, and microRNA. Recent studies showed that EVs are highly correlated with cancer progression and metastasis. However, there are some difficulties in probing each vesicle using popular analytical methods because of their small sizes and heterogeneous origins. These obstacles may be overcome by using a novel approach that senses highly curved membrane and negatively charged membrane lipids. In this chapter, we highlight the basic biological concepts of EVs, isolation, and quantification methods, and recent advent of peptide probes for EVs.

Vitamin D and probiotics supplement use in young children with genetic risk for type 1 diabetes.

BACKGROUND/OBJECTIVES: Vitamin D and probiotics are nutrients of interest in the context of type 1 diabetes (T1D). We assessed the prevalence of and factors associated with vitamin D and probiotic supplementations among young children with genetic risk of T1D. SUBJECTS/METHODS: Use of supplements during the first 2 years of life was collected prospectively from 8674 children in The Environmental Determinants of Diabetes in the Young (TEDDY) study. RESULTS: Single and/or multivitamin/mineral (MVM) supplements were reported by 81% of the children. The majority of participants in Finland, Germany and Sweden (97-99%) and 50% in the United States received vitamin D supplements that were mostly MVMs. Probiotics use varied from 6% in the United States to 60% in Finland and was primarily from probiotics-only preparations. More than 80% of the vitamin D and probiotics supplementation was initiated during infancy, and more than half of the uses lasted longer than a year. Being the first child, longer duration of breastfeeding, born in a later year, older maternal age and higher maternal education level were associated with both vitamin D and probiotics use. Shorter gestational age and mother not smoking during pregnancy were associated with a higher likelihood of probiotics supplementation only. CONCLUSIONS: Vitamin D and probiotics supplementations are popular in children 0-2 years old and are associated with common factors. Data documented here will allow evaluation of the relationship between early childhood dietary intake and the development of islet autoimmunity and progression to T1D.

Neuronal responses to a delayed-response delayed-reward go/nogo task in the monkey posterior insular cortex.

Anatomical connections of the insular cortex suggest its involvement in cognition, emotion, memory, and behavioral manifestation. However, there have been few neurophysiological studies on the insular cortex in primates, in relation to such higher cognitive functions. In the present study, neural activity was recorded from the monkey insular cortex during performance of a delayed-response delayed-reward go/nogo task. In this task, visual stimuli indicating go or nogo responses associated with reward (reward trials) and with no reward (no-reward trials) were presented after eye fixation. In the reward trials, the monkey was required to release a button during presentation of the 2nd visual stimuli after a delay period (delay 1). Then, a juice reward was delivered after another delay (delay 2). The results indicated that the neurons responding in each epoch of the task were topographically localized within the insular cortex, consistent with the previous anatomical studies indicating topographical distributions of afferent inputs from other subcortical and cortical sensory areas. Furthermore, some insular neurons 1) nonspecifically responded to the visual cues and during fixation; 2) responded to the visual cues predicting reward and during the delay period before reward delivery; 3) responded differentially in go/nogo trials during the delay 2; and 4) responded around button manipulation. The observed patterns of insular-neuron responses and the correspondence of their topographical localization to those in previous anatomical studies suggest that the insular cortex is involved in attention- and reward-related functions and might monitor and integrate activities of other brain regions during cognition and behavioral manifestation.

A monoclonal antibody inhibits adhesion to fibronectin and vitronectin of a colon carcinoma cell line and recognizes the integrins alpha v beta 3, alpha v beta 5, and alpha v beta 6.

Using whole viable human colon carcinoma HT29 cells as immunogen, we produced a monoclonal antibody (mAb) termed 69-6-5. The antibody was functionally selected on its anti-cell-spreading activity. By immunoprecipitation of surface radiolabeled cell lysates from HT29-D4 cells (an HT29 cell clone), mAb 69-6-5 recognized a molecular complex resembling integrin heterodimers. Sequential immunodepletions with mAb to the integrin alpha v subunit demonstrated that this complex was composed of alpha v-containing integrins. Accordingly, mAb 69-6-5 reacted with integrin alpha v beta 3 immunopurified from melanoma cells and integrins alpha v beta 5 and alpha v beta 6 immunopurified from pancreatic carcinoma cells. In cell adhesion assays, the 69-6-5 mAb was able to inhibit strongly in a dose-dependent manner arginine-glycine-aspartic acid-mediated adhesion of HT29-D4 cells to vitronectin, fibronectin, or ProNectin F but not to laminin or collagen. Immunoprecipitations with beta chain-specific antisera indicated that these cells express integrins alpha v beta 5 (receptor for vitronectin) and alpha v beta 6 (receptor for fibronectin) but neither alpha v beta 1 nor alpha v beta 3. In summary, these results indicated that mAb 69-6-5 reacts with several alpha v integrins and that it can effectively interfere with the adhesive functions of at least alpha v beta 5 and alpha v beta 6, which represent the major receptors on HT29-D4 cells responsible for their adhesion on vitronectin and fibronectin.

Experimental induction of neoplasia in the accessory sex organs of male Lobund-Wistar rats.

Experimental induction of neoplasia in the urogenital tract was studied in male Lobund-Wistar rats. Animals were given single 30.0-mg/kg i.v. injections of N-nitroso-N-methylurea (NMU) followed 7 days later by s.c. implantation of a 2.0-cm Silastic capsule containing testosterone propionate (TP). Additional rats were given the NMU or TP treatments individually. Control animals were given a single i.v. injection of saline followed by implantation of an empty Silastic capsule. The Silastic implants for each group were replaced every 2 months. This hormone treatment regimen produced significantly (P less than 0.05) elevated serum testosterone concentrations relative to control for 42 days following implantation. Animals were killed at 92, 177, 259, 361, or 427 days post-NMU injection. A high treatment-related incidence of adenocarcinoma occurred in the dorsal and lateral prostatic lobes of animals given the combined NMU-TP treatment. In addition, a few animals had adenocarcinomas of the coagulating gland or the seminal vesicle. The estimated probability of neoplasia in the accessory sex organs by 427 days after initiation of the NMU-TP treatment was 68%, with no occurrence before 9 months. The NMU-TP treatment was also associated with an incidence of focal dysplasia in the accessory sex organs, particularly in the coagulating gland. These findings indicate that NMU-TP treatment of Lobund-Wistar rats can provide a useful experimental system to study the biochemical and molecular events involved in the induction of accessory sex organ neoplasia.

Scoring system to predict hemorrhage in pelvic ring fracture.

BACKGROUND: Risk factors for hemorrhage in patients with pelvic ring fracture have been widely reported. Because there are many risk factors, it is thought that prediction accuracy of hemorrhage in cases of pelvic ring fracture could be improved by using a scoring system. HYPOTHESIS: We investigated the risk factors for massive hemorrhage (MH) and created a novel predictive score of MH in pelvic ring fractures. MATERIAL AND METHODS: We retrospectively reviewed patients with pelvic ring fractures (Abbreviated Injury Score≥3 and age≥16 years) from January 2007 to June 2015. We excluded the cases that might have hemorrhage from other sites sufficient to require a blood transfusion. Massive hemorrhage was defined as hemorrhage requiring transfusion of≥6 red cell concentrate units within 24h of admission. RESULTS: The MH group included 27 patients and the non-MH group included 71 patients. Lactate level, AO/OTA classification and extravasation of computed tomography (CT) contrast fluid had a significantly higher risk as a result of multivariable analysis. The combined score using these risk factors according to their odds-adjusted ratios was created to predict for MH: lactate level>2.5-5.0 (mmol/L)=1 point,>5.0 (mmol/L)=2 points, partially stable (OA/OTA classification B1/B2/B3)=1 point, unstable (C1/C2/C3)=2 points, pelvic extravasation of contrast on CT=4 points. The AUC of the calculated score was 0.93 (95% CI: 0.89-0.98). CONCLUSION: The combined score using these risk factors according to their odds-adjusted ratios was created to predict MH and was an effective prediction score. LEVEL OF EVIDENCE: IV, retrospective study.

Effect of pyrimidine deoxynucleosides and sodium butyrate on expression of the glycoprotein hormone alpha-subunit and placental alkaline phosphatase in HeLa cells.

Production of the glycoprotein hormone common alpha-subunit and placental alkaline phosphatase activity can be modulated in HeLa cells by a variety of deoxynucleosides. Dose response curves for thymidine (Thd), fluorodeoxyuridine (FdUrd), bromodeoxyuridine (BrdUrd) and iododeoxyuridine (IdUrd) demonstrate that, in general, alkaline phosphatase was increased by lower concentrations of inducer than was alpha-subunit. The deoxynucleosides were not as effective as sodium butyrate as inducers of either protein. Whereas Thd and the halogenated dUrd derivatives enhanced protein expression, deoxycytidine (dCyd) had negative effects. Induction by deoxynucleosides of both alkaline phosphatase and alpha-subunit was inhibited by dCyd, but induction of alkaline phosphatase by butyrate was more sensitive to dCyd inhibition than was the butyrate-mediated induction of alpha-subunit. These results suggest that the two proteins are not regulated in a coordinate manner. Reversal of alkaline phosphatase induction by dCyd was not observed in cells preincubated with sodium butyrate for 6-24 h before the addition of dCyd, indicating that the deoxynucleoside interferes with an early event in the butyrate-mediated response. Combinations of butyrate with Thd, BrdUrd or IdUrd were synergistic with respect to the induction of HeLa-alpha. It is concluded that incorporation of the deoxynucleosides into DNA may not be required for the synergistic response since 2',5'-dideoxythymidine was an effective as Thd. Cytoplasmic dot hybridizations demonstrate that a primary effect of the various effectors is to increase the steady-state levels of alpha-subunit mRNA. There was a good correlation between alpha-subunit accumulation and corresponding levels of alpha-mRNA, suggesting that regulation occurs at a pretranslational site. Although the mechanism(s) is not understood, these data provide evidence that nucleosides or their derivatives can significantly affect gene expression.

Immunochemical studies on the subunits of rabbit-intestinal sucrase-isomaltase complex.

Purified sucrase-isomaltase complex sucrose alpha-glucohydrolase, EC 3.2.1.48 - dextrin 6-alpha-glycanohydrolase, EC 3.2.1.10) solubilized by papain from rabbit intestine was dissociated by citraconylation into its subunits, sucrase and isomaltase, which were then isolated in a form active immunologically as well as enzymatically by affinity chromatography on Sephadex G-200 and gel-filtration on Bio-gel P-300. Antibodies against the purified complex inhibited isomaltase but not sucrase and formed precipitation lines, crossing each other, with isolated sucrase and isomaltase, showing that the two enzymes differ in antigenicity from each other. By absorbing the antibodies with isolated sucrase and isomaltase, antibodies specific for isomaltase and sucrase, respectively, were obtained. Like the original antibodies, both of the specific antibodies quantitatively agglutinated microvillous vesicles. Sucrase was inhibited by neither of the antibodies. In contrast, isomaltase was greatly inhibited by the isomaltase-specific antibodies, but not by the sucrase-specific ones.

Immunological similarity between NADH-cytochrome b5 reductase of erythrocytes and liver microsomes.

In a number of animal species soluble NADH-cytochrome b5 reductase of erythrocytes was compared with membrane-bound NADH-cytochrome b5 reductase of liver microsomes by using an antibody to purified NADH-cytochrome b5 reductase from rat liver microsomes. The results obtained indicated clearly that they are immunologically very similar to each other. The data with erythrocyte ghosts suggested that cytochrome b5 and NADH-cytochrome b5 reductase are also present in the ghost.

Olanzapine treatment of psychotic and behavioral symptoms in patients with Alzheimer disease in nursing care facilities: a double-blind, randomized, placebo-controlled trial. The HGEU Study Group.

BACKGROUND: Patients with Alzheimer disease (AD) commonly exhibit psychosis and behavioral disturbances that impair patient functioning, create caregiver distress, and lead to institutionalization. This study was conducted to assess the efficacy and safety of olanzapine in treating psychosis and/or agitation/aggression in patients with AD. METHODS: A multicenter, double-blind, placebo-controlled, 6-week study was conducted in 206 elderly US nursing home residents with AD who exhibited psychotic and/or behavioral symptoms. Patients were randomly assigned to placebo or a fixed dose of 5, 10, or 15 mg/d of olanzapine. The primary efficacy measure was the sum of the Agitation/Aggression, Hallucinations, and Delusions items (Core Total) of the Neuropsychiatric Inventory-Nursing Home version. RESULTS: Low-dose olanzapine (5 and 10 mg/d) produced significant improvement compared with placebo on the Core Total (-7.6 vs -3.7 [P<.001] and -6.1 vs -3. 7 [P =.006], respectively). Core Total improvement with olanzapine, 15 mg/d, was not significantly greater than placebo. The Occupational Disruptiveness score, reflecting the impact of patients' psychosis and behavioral disturbances on the caregiver, was significantly reduced in the 5-mg/d olanzapine group compared with placebo (-2.7 vs -1.5; P =.008). Somnolence was significantly more common among patients receiving olanzapine (25.0%-35.8%), and gait disturbance occurred in those receiving 5 or 15 mg/d (19.6% and 17.0%, respectively). No significant cognitive impairment, increase in extrapyramidal symptoms, or central anticholinergic effects were found at any olanzapine dose relative to placebo. CONCLUSION: Low-dose olanzapine (5 and 10 mg/d) was significantly superior to placebo and well tolerated in treating agitation/aggression and psychosis in this population of patients with AD.

p19Arf sensitizes B16 melanoma cells to interferon-β delivered via mesenchymal stem cells in vitro

The immune stimulatory and anti-neoplastic functions of type I interferon have long been applied for the treatment of melanoma. However, the systemic application of high levels of this recombinant protein is often met with toxicity. An approach that provides localized, yet transient, production of type I interferon may overcome this limitation. We propose that the use of mesenchymal stem cells (MSCs) as delivery vehicles for the production of interferon-β (IFNβ) may be beneficial when applied together with our cancer gene therapy approach. In our previous studies, we have shown that adenovirus-mediated gene therapy with IFNβ was especially effective in combination with p19Arf gene transfer, resulting in immunogenic cell death. Here we showed that MSCs derived from mouse adipose tissue were susceptible to transduction with adenovirus, expressed the transgene reliably, and yet were not especially sensitive to IFNβ production. MSCs used to produce IFNβ inhibited B16 mouse melanoma cells in a co-culture assay. Moreover, the presence of p19Arf in the B16 cells sensitizes them to the IFNβ produced by the MSCs. These data represent a critical demonstration of the use of MSCs as carriers of adenovirus encoding IFNβ and applied as an anti-cancer strategy in combination with p19Arf gene therapy.

Rapid spreading and mature hemidesmosome formation in HaCaT keratinocytes induced by incubation with soluble laminin-5r.

HaCaT cells, an immortalized keratinocyte line, incubated in plastic wells in the presence of conditioned medium from 804G cells adhered and spread rapidly in less than 30 min. In contrast, cells plated in fibroblast or keratinocyte conditioned medium adhered poorly and remained rounded at 30 min. Immunodepletion of 804G conditioned medium with polyclonal antisera to laminin-5r, but not control antisera, abolished rapid cell spreading. Electron microscopy of HaCaT cells spread by incubation in 804G conditioned medium, but not control medium, revealed mature hemidesmosomes after 24 h. Rapid spreading was also observed in wells precoated with 804G conditioned medium or 804G cell-deposited matrix, but not with fibronectin, vitronectin, or laminin-1. Immunoblotting of 804G conditioned medium with anti-laminin-5r antibodies unveiled polypeptides of 150, 140, 135, and 100 kDa, identical by electrophoretic mobility to immunoreactive polypeptides in 804G deposited matrix. Our results suggest that addition of laminin-5r in a soluble form is sufficient to promote rapid spreading and hemidesmosome assembly in keratinocytes. The mechanism of soluble laminin-5r action may include efficient surface "priming" for cell adhesion. Soluble laminin-5r may have a physiologic role in morphogenesis and repair of the epidermis and may be of use for therapeutic applications.

Blind, controlled, long-term study of the comparative incidence of treatment-emergent tardive dyskinesia with olanzapine or haloperidol.

OBJECTIVE: Tardive dyskinesia is a serious and common complication of neuroleptic treatment. Olanzapine is a novel antipsychotic agent exhibiting regional mesolimbic dopaminergic selectivity and a broad-based pharmacology encompassing serotonin, dopamine, muscarinic, and adrenergic receptor binding affinities. The authors' goal was to compare the incidence of tardive dyskinesia among patients receiving olanzapine and those receiving the conventional dopamine 2 antagonist haloperidol. METHOD: Data were analyzed from three actively controlled and blind long-term responder studies of subjects meeting DSM-III-R criteria for schizophrenia, schizophreniform disorder, or schizoaffective disorder treated with olanzapine (N = 707, up to 20 mg/day, 237 median days of exposure) or haloperidol (N = 197, up to 20 mg/day, 203 median days of exposure) who did not have evidence of tardive dyskinesia at baseline. All of the subjects had a chronic disease course (mean greater than 10 years), and there were no significant between-treatment group differences in demographic or disease characteristics. The Abnormal Involuntary Movement Scale and research diagnostic criteria for tardive dyskinesia were used to define the comparative incidence rates of long-term treatment-emergent tardive dyskinesia. RESULTS: The incidence of newly emergent tardive dyskinesia at any visit after baseline, at the final visit, and at the final two clinical assessments was statistically significantly lower among olanzapine-treated patients than among haloperidol-treated patients. CONCLUSIONS: These findings support an atypical extrapyramidal symptom profile and the potential of a significantly lower risk of tardive dyskinesia with olanzapine than with haloperidol among patients requiring maintenance antipsychotic treatment.