Serially transplantable mammary epithelial cells express the Thy-1 antigen

Abstract Background Recent studies in murine mammary tissue have identified functionally distinct cell populations that may be isolated by surface phenotype or lineage tracing. Previous groups have shown that CD24medCD49fhigh cells enriched for long-lived mammary epithelial cells can be serially transplanted. Methods Flow cytometry-based enrichment of distinct phenotypic populations was assessed for their gene expression profiles and functional proliferative attributes in vitro and in vivo. Results Here, we show Thy-1 is differentially expressed in the CD24medCD49fhigh population, which allowed us to discern two functionally different populations. The Thy-1+CD24medCD49fhigh phenotype contained the majority of the serially transplantable epithelial cells. The Thy-1−CD24medCD49fhigh phenotype contains a rare progenitor population that is able to form primary mammary outgrowths with significantly decreased serial in vivo transplantation potential. Conclusions Therefore, Thy-1 expression in the immature cell compartment is a useful tool to study the functional heterogeneity that drives mammary gland development and has implications for disease etiology.https://deepblue.lib.umich.edu/bitstream/2027.42/145730/1/13058_2018_Article_1006.pd

New Oncolytic Adenoviruses with Hypoxia- and Estrogen Receptor-Regulated Replication

Oncolytic adenoviruses with restricted replication can be produced if the expression of crucial transcription units of the virus is controlled by tissue- or tumor-specific promoters. Here we describe a method for the rapid incorporation of exogenous promoters into the E1A and E4 regions of the human adenovirus type 5 genome. Using this system, we have generated AdEHT2 and AdEHE2F, two conditionally replicative adenoviruses for the treatment of breast cancer. The expression of the E1A gene in both viruses is controlled by a minimal dual-specificity promoter that responds to estrogens and hypoxia. The tight regulation of E1A expression correlated with the ability of these viruses to replicate and kill human cancer cells that express estrogen receptors, or are maintained under hypoxic conditions. The telomerase reverse transcriptase (TERT) promoter and the E2F-1 promoter are preferentially activated in cancer cells. They were introduced into the E4 region of AdEHT2 and AdEHE2F, respectively. The telomerase core promoter failed to block the replication of the virus in telomerase-negative cells. In contrast, AdEHE2F was attenuated in nontransformed quiescent cells growing under normoxic conditions, suggesting that an intact pRB pathway with low levels of E2F transcription factors acts as a negative modulator for the virus. These data indicate that the simultaneous regulation of E1A and E4 viral transcription units by the appropriate combination of promoters can increase the tumor selectivity of oncolytic adenoviruses.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/63195/1/104303402760293574.pd

A Method of Limited Replication for the Efficient In Vivo Delivery of Adenovirus to Cancer Cells

Overview summary Replication-defective viral vectors are limited in their ability to diffuse through tissue. This poses a problem for treating tumors in vivo using gene transfer. This article demonstrates that limited replication of adenovirus leads to greater gene transfer efficiency in vitro and in vivo without introducing additional safety concerns beyond traditional adenovirus administration. This has implications for the improvement of current gene transfer methods for treating cancer.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/63280/1/hum.1998.9.8-1209.pd

Bmi-1 is required for maintenance of adult self-renewing haematopoietic stem cells

A central issue in stem cell biology is to understand the mechanisms that regulate the self-renewal of haematopoietic stem cells (HSCs), which are required for haematopoiesis to persist for the lifetime of the animal(1). We found that adult and fetal mouse and adult human HSCs express the proto-oncogene Bmi-1. The number of HSCs in the fetal liver of Bmi-1(-/-) mice(2) was normal. In postnatal Bmi-1(-/-) mice, the number of HSCs was markedly reduced. Transplanted fetal liver and bone marrow cells obtained from Bmi-1(-/-) mice were able to contribute only transiently to haematopoiesis. There was no detectable self-renewal of adult HSCs, indicating a cell autonomous defect in Bmi-1(-/-) mice. A gene expression analysis revealed that the expression of stem cell associated genes(3), cell survival genes, transcription factors, and genes modulating proliferation including p16(Ink4a) and p19(Arf) was altered in bone marrow cells of the Bmi-1(-/-) mice. Expression of p16(Ink4a) and p19(Arf) in normal HSCs resulted in proliferative arrest and p53-dependent cell death, respectively. Our results indicate that Bmi-1 is essential for the generation of self-renewing adult HSCs.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/62508/1/nature01587.pd

Dynamic Alterations in Spontaneous Neural Activity in Multiple Brain Networks in Subacute Stroke Patients: A Resting-State fMRI Study

Objective: To examine whether subacute stroke patients would exhibit abnormal dynamic characteristics of brain activity relative to healthy controls (HC) and to investigate whether the altered dynamic regional indexes were associated with clinical behavior in stroke patients.Methods: The dynamic amplitude of low-frequency fluctuations (dALFF) and dynamic regional homogeneity (dReHo) in 42 subacute stroke patients and 55 healthy controls were compared. Correlation analyses between dALFF and dReHo in regions showing significant intergroup differences and clinical scores (i.e., the National Institutes of Health Stroke Scale, Fugl-Meyer assessment and lesion volume size) were conducted in stroke patients. Receiver operating characteristic (ROC) curve analysis was used to determine the potential value of altered dynamic regional indexes to identify stroke patients.Results: Significantly dALFF in the bilateral cerebellum posterior lobe (CPL), ipsilesional superior parietal lobe, ipsilesional inferior temporal gyrus (ITG), the midline supplementary motor area (SMA), ipsilesional putamen and lentiform nucleus were detected in stroke patients compared to HC. Relative to the HC group, the stroke patients showed significant differences in dReHo in the contralesional rectal gyrus, contralesional ITG, contralesional pons, ipsilesional middle frontal gyrus (MFG). Significant correlations between dALFF variability in midline SMA and Fugl-Meyer assessment (FMA) scores or between dReHo variability in the ipsilesional MFG and FMA scores were detected in stroke patients. Furthermore, the ROC curve revealed that dynamic ALFF at SMA and ReHo at ipsilesional MFG might have the potential to distinguish stroke patients.Conclusion: The pattern of intrinsic brain activity variability is altered in stroke patients compared with HC, and dynamic ALFF/ReHo might be potential tools to assess stroke patients’ motor function

Picturing Electron Capture to the Continuum in the Transfer Ionization of Intermediate-Energy He²⁺ Collisions with Argon

Electron emission occurring in transfer ionization for He2+ collisions with argon has been investigated using cold target recoil ion momentum spectroscopy. The double differential cross sections for electron capture to the continuum of the projectile (cusp-shaped electrons) are presented for collision energies from 17.5 to 75 keV/u. For an energy of 30 keV/u, we find a maximum in the experimental ratio of the cusp-shaped electron yield to the total electron yield. This result is explained in terms of the velocity matching between the projectile ion and the electron initially bound to the target. One of the important issues for double electron transitions is the role of electron-electron correlation. If this correlation is weak, then the transfer-ionization process can be viewed as two separate sequential processes. If this correlation is strong, then the transfer-ionization process would happen simultaneously and not sequentially. Our experimental and theoretical results indicate that correlation is weak and that the first step is target ionization followed by charge capture

CDX2 as a Prognostic Biomarker in Stage II and Stage III Colon Cancer

BACKGROUND: The identification of high-risk stage II colon cancers is key to the selection of patients who require adjuvant treatment after surgery. Microarray-based multigene-expression signatures derived from stem cells and progenitor cells hold promise, but they are difficult to use in clinical practice. METHODS: We used a new bioinformatics approach to search for biomarkers of colon epithelial differentiation across gene-expression arrays and then ranked candidate genes according to the availability of clinical-grade diagnostic assays. With the use of subgroup analysis involving independent and retrospective cohorts of patients with stage II or stage III colon cancer, the top candidate gene was tested for its association with disease-free survival and a benefit from adjuvant chemotherapy. RESULTS: The transcription factor CDX2 ranked first in our screening test. A group of 87 of 2115 tumor samples (4.1%) lacked CDX2 expression. In the discovery data set, which included 466 patients, the rate of 5-year disease-free survival was lower among the 32 patients (6.9%) with CDX2-negative colon cancers than among the 434 (93.1%) with CDX2-positive colon cancers (hazard ratio for disease recurrence, 3.44; 95% confidence interval [CI], 1.60 to 7.38; P = 0.002). In the validation data set, which included 314 patients, the rate of 5-year disease-free survival was lower among the 38 patients (12.1%) with CDX2 protein–negative colon cancers than among the 276 (87.9%) with CDX2 protein–positive colon cancers (hazard ratio, 2.42; 95% CI, 1.36 to 4.29; P = 0.003). In both these groups, these findings were independent of the patient's age, sex, and tumor stage and grade. Among patients with stage II cancer, the difference in 5-year disease-free survival was significant both in the discovery data set (49% among 15 patients with CDX2-negative tumors vs. 87% among 191 patients with CDX2-positive tumors, P = 0.003) and in the validation data set (51% among 15 patients with CDX2-negative tumors vs. 80% among 106 patients with CDX2-positive tumors, P = 0.004). In a pooled database of all patient cohorts, the rate of 5-year disease-free survival was higher among 23 patients with stage II CDX2-negative tumors who were treated with adjuvant chemotherapy than among 25 who were not treated with adjuvant chemotherapy (91% vs. 56%, P = 0.006). CONCLUSIONS: Lack of CDX2 expression identified a subgroup of patients with high-risk stage II colon cancer who appeared to benefit from adjuvant chemotherapy. (Funded by the National Comprehensive Cancer Network, the National Institutes of Health, and others.

Effects of bacterial endotoxin on liver mass and hepatocyte volume

Hepatomegaly is common in infections. An increase in liver mass, which cannot be accounted for by the net increase of hepatic protein and lipid contents, has been reported to be a common tumour necrosis factor a (TNFα)- mediated host response after such stress stimuli as endotoxemia, sepsis, trauma and chronic inflammation in either fasted or fed rats. The increase of hepatocyte volume has also been associated with, in experiments in vitro, alterations in hepatic metabolic functions such as protein and carbohydrate metabolism. There are, however, no previous studies carried out on alterations in liver mass and hepatocyte volume in vivo as a host response of endotoxic animals which undergo profound metabolic changes in liver. -- In this study, a single intraperitoneal injection of E. coli lipopolysaccharide (LPS, 0127:B8) at a sublethal dose (3 mg/kg, LD₅) was found to introduce a generally time- and dose-dependent increase in hepatic mass and water content of fasted rats compared to corresponding saline-injected controls. Twenty-four hr after LPS treatment, the absolute and relative weight of livers removed by complete hepatectomy remain unaltered in fasted endotoxic rats but they decrease by 20% in the fasted saline-controls, compared to that in the normal rats fed ad libitum. The difference in the mean liver weight between the endotoxin- and saline-treated animals is 2.0-2.5 g, i.e. 20-25% of the total liver mass of 300-350 g rats. The hepatic wet/dry weight ratio is significantly higher in endotoxic livers than that in the other two groups. The hepatic water content increases by almost the same extent (27%) as the increase (26%) in the total liver mass 24 hr after treatment, indicating that the effect of E. coli LPS is mainly on the hepatic water content, rather than on the dry mass components. -- Analyses of hepatic dry mass components and hepatic cellular spaces show that only one third (0.79 g) of the increase (2.18 g) in the total liver mass can be accounted for by the net increase of the dry mass components (0.38 g) plus the blood/plasma and/or interstitial fluid (0.41 ml) in the extrahepatocellular space; and hepatic K⁺ content, the predominant intracellular cation, increases by 25%, 24 hr after LPS injection. These suggest that an expansion of the hepatocellular space is mainly responsible for the increase in the liver mass of endotoxic rats. This hypothesis is also supported by morphometric data from an image analysis system demonstrating that the mean transsectional area and the mean volume is 8% and 12% larger, respectively, in the mononucleate hepatocytes from the endotoxic livers. -- The phenomenon of increases in the hepatic mass, water content and wet/dry weight ratio after LPS treatment can be reproduced by a single intravenous administration of rhTNF-α (25 μg/250 g B.W.) and prevented by the pretreatment of endotoxic rats with an intraperitoneal injection of goat anti-mTNFα serum (20 mg/300 g B.W.). The mean transsectional area and mean volume is 11% and 17% larger, respectively, in mononucleate hepatocytes from the livers of endotoxic rats treated with goat non-immune IgG compared to the anti-mTNFα serum-treated rats. These data suggest that effects of E. coli LPS on the increase in hepatic mass and hepatocyte volume are mediated by cytokine TNF-α. -- Twenty-four hr after E. coli LPS injection, the concentrations of hepatic adenine nucleotides in the endotoxic rats are not significantly different from either the saline-controls or the normal fed rats. There are no significant changes in the activity of SGPT or in the plasma ratio of lactate/pyruvate and β-hydroxybutyrate/acetoacetate during the 24 hr after LPS injection. Those observations argue against "compensatory hyperplasia" resultinq from acute liver damage due to direct endotoxin toxicity or hepatocyte swelling due to tissue hypoxia and impaired energy status in the liver. Metabolic changes mediated by insulin, TNF-α and/or other "hepatotrophic factors" may be responsible for the increase of liver mass after E. coli LPS administration