Residual embryonic cells as precursors of a Barrett's-like metaplasia

Barrett's esophagus is an intestine-like metaplasia and precursor of esophageal adenocarcinoma. Triggered by gastroesophageal reflux disease, the origin of this metaplasia remains unknown. p63-deficient mice, which lack squamous epithelia, may model acid-reflux damage. We show here that p63 null embryos rapidly develop intestine-like metaplasia with gene expression profiles similar to Barrett's metaplasia. We track its source to a unique embryonic epithelium that is normally undermined and replaced by p63-expressing cells. Significantly, we show that a discrete population of these embryonic cells persists in adult mice and humans at the squamocolumnar junction, the source of Barrett's metaplasia. We show that upon programmed damage to the squamous epithelium, these embryonic cells migrate toward adjacent, specialized squamous cells in a process that may recapitulate early Barrett's. Our findings suggest that certain precancerous lesions, such as Barrett's, initiate not from genetic alterations but from competitive interactions between cell lineages driven by opportunity.     

Regulation of ABCC6 Trafficking and Stability by a Conserved C-terminal PDZ-Like Sequence

Mutations in the ABCC6 ABC-transporter are causative of pseudoxanthoma elasticum (PXE). The loss of functional ABCC6 protein in the basolateral membrane of the kidney and liver is putatively associated with altered secretion of a circulatory factor. As a result, systemic changes in elastic tissues are caused by progressive mineralization and degradation of elastic fibers. Premature arteriosclerosis, loss of skin and vascular tone, and a progressive loss of vision result from this ectopic mineralization. However, the identity of the circulatory factor and the specific role of ABCC6 in disease pathophysiology are not known. Though recessive loss-of-function alleles are associated with alterations in ABCC6 expression and function, the molecular pathologies associated with the majority of PXE-causing mutations are also not known. Sequence analysis of orthologous ABCC6 proteins indicates the C-terminal sequences are highly conserved and share high similarity to the PDZ sequences found in other ABCC subfamily members. Genetic testing of PXE patients suggests that at least one disease-causing mutation is located in a PDZ-like sequence at the extreme C-terminus of the ABCC6 protein. To evaluate the role of this C-terminal sequence in the biosynthesis and trafficking of ABCC6, a series of mutations were utilized to probe changes in ABCC6 biosynthesis, membrane stability and turnover. Removal of this PDZ-like sequence resulted in decreased steady-state ABCC6 levels, decreased cell surface expression and stability, and mislocalization of the ABCC6 protein in polarized cells. These data suggest that the conserved, PDZ-like sequence promotes the proper biosynthesis and trafficking of the ABCC6 protein.     

Effect of Dietary Starch Source and Concentration on Equine Fecal Microbiota

Starch from corn is less susceptible to equine small intestinal digestion than starch from oats, and starch that reaches the hindgut can be utilized by the microbiota. The objective of the current study was to examine the effects of starch source on equine fecal microbiota. Thirty horses were assigned to treatments: control (hay only), HC (high corn), HO (high oats), LC (low corn), LO (low oats), and LW (low pelleted wheat middlings). Horses received an all-forage diet (2 wk; d -14 to d -1) before the treatment diets (2 wk; d 1 to 14). Starch was introduced gradually so that horses received 50% of the assigned starch amount (high = 2 g starch/kg BW; low = 1 g starch/kg BW) by d 4 and 100% by d 11. Fecal samples were obtained at the end of the forage-only period (S0; d -2), and on d 6 (S1) and d 13 (S2) of the treatment period. Cellulolytics, lactobacilli, Group D Gram-positive cocci (GPC), lactate-utilizers and amylolytics were enumerated. Enumeration data were log transformed and analyzed by repeated measures ANOVA. There were sample day × treatment interactions (P < 0.0001) for all bacteria enumerated. Enumerations from control horses did not change during the sampling period (P > 0.05). All treatments except LO resulted in increased amylolytics and decreased cellulolytics, but the changes were larger in horses fed corn and wheat middlings (P < 0.05). Feeding oats resulted in increased lactobacilli and decreased GPC (P < 0.05), while corn had the opposite effects. LW had increased lactobacilli and GPC (P < 0.05). The predominant amylolytic isolates from HC, LC and LW on S2 were identified by 16S RNA gene sequencing as Enterococcus faecalis, but other species were found in oat fed horses. These results demonstrate that starch source can have a differential effect on the equine fecal microbiota.     

Beta-catenin accelerates human papilloma virus type-16 mediated cervical carcinogenesis in transgenic mice

Human papilloma virus (HPV) is the principal etiological agent of cervical cancer in women, and its DNA is present in virtually all of these tumors. However, exposure to the high-risk HPV types alone is insufficient for tumor development. Identifying specific collaborating factors that will lead to cervical cancer remains an unanswered question, especially because millions of women are exposed to HPV. Our earlier work using an in vitro model indicated that activation of the canonical Wnt pathway in HPV-positive epithelial cells was sufficient to induce anchorage independent growth. We therefore hypothesized that constitutive activation of this pathway might function as the "second hit." To address this possibility, we developed two double-transgenic (DT) mouse models, K14-E7/ΔN87βcat and K14-HPV16/ΔN87βcat that express either the proteins encoded by the E7 oncogene or the HPV16 early region along with constitutively active β-catenin, which was expressed by linking it to the keratin-14 (K14) promoter. We initiated tumor formation by treating all groups with estrogen for six months. Invasive cervical cancer was observed in 11% of the K14-ΔN87βcat mice, expressing activated β-catenin and in 50% of the animals expressing the HPV16 E7 oncogene. In double-transgenic mice, coexpression of β-catenin and HPV16 E7 induced invasive cervical cancer at about 7 months in 94% of the cases. We did not observe cervical cancer in any group unless the mice were treated with estrogen. In the second model, K14-HPV16 mice suffered cervical dysplasias, but this phenotype was not augmented in HPV16/ΔN87βcat mice. In summary, the phenotypes of the K14-E7/ΔN87βcat mice support the hypothesis that activation of the Wnt/β-catenin pathway in HPV-associated premalignant lesions plays a functional role in accelerating cervical carcinogenesis.     

Activation of morphogenesis and proliferation by low specificity chemical effectors

Activation of highly specific biochemical processes by simple chemical agents is demonstrated for morphogenesis (anlage and development of female gametophyte in cereal) and mitosis (in cell cultures and animal and plant tissues). The effects of these agents are tissue-specific. Structure--activity relationship is analyzed in this group of compounds. Thus, the phenomenon reveals the exact pathways of the influence of allelopathic and anthropogenic chemical agents on evolution of plant biocenoses.     

Peritoneal Dissemination Complicating Morcellation of Uterine Mesenchymal Neoplasms

Background

Power morcellation has become a common technique for the minimally invasive resection of uterine leiomyomas. This technique is associated with dissemination of cellular material throughout the peritoneum. When morcellated uterine tumors are unexpectedly found to be leiomyosarcomas or tumors with atypical features (atypical leiomyoma, smooth muscle tumor of uncertain malignant potential), there may be significant clinical consequences. This study was undertaken to determine the frequency and clinical consequence of intraperitoneal dissemination of these neoplasms.

Methodology/Principal Findings

From 2005–2010, 1091 instances of uterine morcellation were identified at BWH. Unexpected diagnoses of leiomyoma variants or atypical and malignant smooth muscle tumors occurred in 1.2% of cases using power morcellation for uterine masses clinically presumed to be “fibroids” over this period, including one endometrial stromal sarcoma (ESS), one cellular leiomyoma (CL), six atypical leiomyomas (AL), three smooth muscle tumor of uncertain malignant potential (STUMPs), and one leiomyosarcoma (LMS). The rate of unexpected sarcoma after the laparoscopic morcellation procedure was 0.09%, 9-fold higher than the rate currently quoted to patients during pre-procedure briefing, and this rate may increase over time as diagnostically challenging or under-sampled tumors manifest their biological potential. Furthermore, when examining follow-up laparoscopies, both from in-house and consultation cases, disseminated disease occurred in 64.3% of all tumors (zero of one ESS, one of one CL, zero of one AL, four of four STUMPs, and four of seven LMS). Only disseminated leiomyosarcoma, however, was associated with mortality. Procedures are proposed for pathologic evaluation of morcellation specimens and associated follow-up specimens.

Conclusions/Significance

While additional study is warranted, these data suggest uterine morcellation carries a risk of disseminating unexpected malignancy with apparent associated increase in mortality much higher than appreciated currently.     

Evolution of extra-nigral damage predicts behavioural deficits in a rat proteasome inhibitor model of Parkinson's disease

Establishing the neurological basis of behavioural dysfunction is key to provide a better understanding of Parkinson's disease (PD) and facilitate development of effective novel therapies. For this, the relationships between longitudinal structural brain changes associated with motor behaviour were determined in a rat model of PD and validated by post-mortem immunohistochemistry. Rats bearing a nigrostriatal lesion induced by infusion of the proteasome inhibitor lactacystin into the left-medial forebrain bundle and saline-injected controls underwent magnetic resonance imaging (MRI) at baseline (prior to surgery) and 1, 3 and 5 weeks post-surgery with concomitant motor assessments consisting of forelimb grip strength, accelerating rotarod, and apormorphine-induced rotation. Lactacystin-injected rats developed early motor deficits alongside decreased ipsilateral cortical volumes, specifically thinning of the primary motor (M1) and somatosensory cortices and lateral ventricle hypertrophy (as determined by manual segmentation and deformation-based morphometry). Although sustained, motor dysfunction and nigrostriatal damage were maximal by 1 week post-surgery. Additional volume decreases in the ipsilateral ventral midbrain; corpus striatum and thalamus were only evident by week 3 and 5. Whilst cortical MRI volume changes best predicted the degree of motor impairment, post-mortem tyrosine hydroxylase immunoreactivity in the striatum was a better predictor of motor behaviour overall, with the notable exception of performance in the accelerating rotarod, in which, M1 cortical thickness remained the best predictor. These results highlight the importance of identifying extra-nigral regions of damage that impact on behavioural dysfunction from damage to the nigrostriatal system.     

Epizootiology of an epizootic hemorrhagic disease outbreak in West Virginia

An outbreak of epizootic hemorrhagic disease virus, serotype 2 (EHDV-2) was responsible for localized white-tailed deer (Odocoileus virginianus) mortality in Hardy and Hampshire counties, West Virginia (USA), in the summer and fall of 1993. Using available historical data on regional herd immunity, data opportunistically collected during the epizootic, and postepizootic sampling of hunter-harvested deer, we grossly estimate certain epidemiologic parameters and compare findings to a hypothesis about hemorrhagic disease outbreaks in the Appalachian Mountains. During the epizootic, 57.9 km(2) were actively searched and 228 dead deer were found. Epizootic hemorrhagic disease virus, serotype 2 was isolated from seven of nine deer sampled in Hardy and Hampshire counties. Preepizootic exposure of deer to EHD viruses was unknown, but available data suggest that it was negligible. The geographic distribution of the outbreak was defined by plotting the locations of dead deer found during the outbreak, as well as the locations of deer harvested by hunters after the outbreak that had antibodies to EHDV-2 on a map sectioned into 16.65 km(2) rectangular sections. Sections that included one or more dead deer or hunter-harvested deer with antibodies to EHDV-2 were included in the defined outbreak area. Postoutbreak sampling revealed monospecific EHDV-2 antibodies in 12% of deer harvested by hunters within the defined outbreak area. Based on the available data and accepting certain assumptions, gross calculations suggest that this outbreak appears to have been isolated and probably killed a high percentage of the deer that were infected. This is consistent with the hypothesis that sporadic hemorrhagic disease outbreaks in the Appalachian Mountains are usually localized and severe.     

Isolation of Stable Hemolysin and Catalase Variants of Staphylococcus aureus S6C Includes One with an Exoprotein-Deficient Phenotype

Previous studies suggested that the exoprotein-deficient phenotype of a Δ1058::Tn551 insertion/deletion mutant of Staphylococcus aureus S6C was not owing to the insertion/deletion event, but instead was owing to the inherent instability of the agrC gene during transduction of the Δ1058::Tn551 region into S6C. The purpose of the following study was to examine S6C as a potential source of exoprotein-deficient mutants that would account for their appearance after transposition and transduction. Four stable variants of S6C were isolated that differed in their hemolysin and catalase activities. Surprisingly, the agr regulatory molecule, RNAIII, was undetectable in one of these variants, which most likely accounted for the exoprotein-deficient phenotype of this variant. When the original Δ1058::Tn551 mutation was transduced into the hemolytic, catalase-positive variant of S6C, none of the transductants exhibited an exoprotein-deficient phenotype. These data suggest that, while the exoprotein-deficient phenotype of the S6C variant is most likely due to mutations in the agr regulatory system, these mutations are not caused by the transduction of the Δ1058::Tn551 region into S6C, but instead already exist in an exoprotein-deficient variant of S6C. Received: 6 November 2000 / Accepted: 16 January 2001     

Angiogenesis is stimulated by a tumor-derived endothelial cell growth factor

A growth factor mitogenic for BALB/C 3T3 cells and capillary endothelial cells was isolated from a rat chondrosarcoma and purified to homogeneity. Purification was accomplished by a combination of BioRex 70 cation exchange chromatography and heparin affinity chromatography. The pure chondrosarcoma-derived growth factor (ChDGF) had a molecular weight of about 18,000. The angiogenesis activity of pure ChDGF was tested by measuring its ability to vascularize the chorioallantoic membrane (CAM) and yolk sac membrane of the developing chick. The ability of ChDGF to induce the growth of limbal vessels in the rat cornea was also measured. To quantitate the angiogenesis response, a unit system based on the growth factor activity of ChDGF for 3T3 cells was adopted. ChDGF was found to have a specific activity of about 5 units/ng when applied to 3T3 cells. About 300-600 units of ChDGF in the two types of developing chick membrane and 30-5 units of ChDGF in the rat cornea were found to stimulate noninflammatory angiogenesis.