Obesity-induced Ly6C and Ly6C monocyte subset changes abolish post-ischemic limb conditioning benefits in stroke recovery.

Remote limb conditioning (RLC), performed by intermittent interruption of blood flow to a limb, triggers endogenous tolerance mechanisms and improves stroke outcomes. The underlying mechanism for the protective effect involves a shift of circulating monocytes to a Ly6C proinflammatory subset in normal metabolic conditions. The current study investigates the effect of RLC on stroke outcomes in subjects with obesity, a vascular comorbidity.

Endothelial cell CD36 mediates stroke-induced brain injury via BBB dysfunction and monocyte infiltration in normal and obese conditions.

CD36 expressed in multiple cell types regulates inflammation, vascular function, and innate immunity. Specifically, CD36 in microvascular endothelial cells (ECs) signals to elicit inflammation and causes EC death. This study investigated roles for EC-CD36 on acute stroke pathology in normal and obese conditions.

Chick Embryonic Primary Astrocyte Cultures Provide an Effective and Scalable Model for Authentic Research in a Laboratory Class.

Cell culture provides an impactful tool for undergraduates to study a range of neurobiological processes. While immortalized or cancer cell lines offer a level of convenience for undergraduate research, particularly for larger scale course-based undergraduate research experiences (CUREs) or project-based learning (PBL), primary cell cultures more closely retain the characteristics of the tissue of origin, allowing students to engage in a wider range of authentic research projects. Astrocytes have gained increasing attention for their role in modulating neuronal viability and are at the forefront of neuroprotection research.

The development of painting probes for dual-color and multiple chromosome analysis in the mouse.

The recent development of mouse chromosome painting probes for fluorescence in situ hybridization has extended the use of this common laboratory mammal in cytogenetics. We now report the development of additional painting probes by degenerate-oligonucleotide-primed PCR on chromosomes from mouse lung fibroblast cultures, each homozygous for a single Robertsonian translocation chromosome. These probes are for Rb(1.

The development of chromosome-specific composite DNA probes for the mouse and their application to chromosome painting.

The speed and ease of human cytogenetic analysis has been greatly enhanced by the technique of fluorescence in situ hybridization (FISH). Non-radioactive fluorescently tagged complex DNA probes specific for individual chromosomes can be hybridized to conventionally obtained metaphase chromosome spreads. Several chromosomes may be "painted" concurrently by using combinations of different labeled probes.

Validation of chromosome painting as a biodosimeter in human peripheral lymphocytes following acute exposure to ionizing radiation in vitro.

Fluorescence in situ hybridization with chromosome-specific composite DNA probes ('chromosome painting') appears to be a useful tool for quantifying symmetrical cytogenetic damage. However, a thorough comparison between chromosome painting and the conventional methods of GTG-banding and dicentric analysis has not been performed. We have undertaken the validation of chromosome painting using human blood exposed in vitro to 137Cs gamma-rays at doses ranging from 0 to 400 cGy, then cultured according to standard procedures and harvested at 52 h.

Biological effectiveness of neutrons from Hiroshima bomb replica: results of a collaborative cytogenetic study.

The effectiveness of neutrons from a facsimile of the Hiroshima bomb was determined cytogenetically. The "Little-Boy" replica (LBR), assembled at Los Alamos as a controlled nuclear reactor for detailed physical dosimetry, was used. Of special interest, the neutron energy characteristics (including lineal energy) measured 0.

Metabolic activation and cytogenetic effects of 2-amino-1-methyl-6-phenylimidazo[4,5-b] pyridine (PhIP) in Chinese hamster ovary cells expressing murine cytochrome P450 IA2.

We have utilized Chinese hamster ovary cell lines which stably express a murine cytochrome P450IA2 (P(3)450) cDNA to characterize more fully the mechanisms of genotoxicity of heterocyclic amines derived from cooked meats. To verify that these cell lines were capable of converting promutagens into active metabolites, we studied the microsomal metabolism and cytogenetic effects of 2-amino-1-methyl-6-phenylimidazo-[4,5-b]pridine (PhIP). Microsomal preparations derived from excision repair-deficient Chinese hamster ovary cells expressing the mouse cytochrome P(3)450 cDNA (UV5P3) converted PhIP to the genotoxic N-hydroxy-PhIP metabolite.

Cytogenetic characterization of the ionizing radiation-sensitive Chinese hamster mutant irs1.

The X-ray-sensitive mutant V79 cell line irs1 was characterized with respect to chromosomal aberrations induced by 137Cs, mitomycin C (MMC), and decarbamoyl mitomycin C (DCMMC). To measure chromosome damage induced at different cell cycle stages, irs1 and the parental V79-4 cell lines were pulse-labeled with bromodeoxyuridine (BrdUrd) at the time of exposure and harvested at various intervals corresponding to exposure in G1, S, and G2 phases of the cell cycle. Metaphase spreads were stained with an anti-BrdUrd antibody, followed by a fluorescein-conjugated second antibody.

Identification of nucleotide-excision-repair genes on human chromosomes 2 and 13 by functional complementation in hamster-human hybrids.

The CHO UV-sensitive mutants UV24 and UV135 (complementation groups 3 and 5, respectively) are defective in nucleotide excision repair. After fusing each mutant with human lymphocytes, resistant hybrid clones showing genetic complementation were isolated by repeated exposure to UV radiation. Using a combination of isozyme markers, DNA probes, and cytogenetic methods to analyze the primary hybrids and their subclones, correction of the repair defect was shown to be correlated with the presence of a specific human chromosome in each case.

Incorporated bromodeoxyuridine enhances the sister-chromatid exchange and chromosomal aberration frequencies in an EMS-sensitive Chinese hamster cell line.

The mutant Chinese hamster cell line, EM9, is characterized by a high baseline sister-chromatid exchange (SCE) frequency, increased sensitivity to cell killing, and a defect in DNA strand-break repair. The molecular basis for this pleotrophic phenotype is not known. We examined, at the chromosomal level, the increased sensitivity of this mutant to incorporated BrdUrd.

DNA-mediated transfer of a human DNA repair gene that controls sister chromatid exchange.

The Chinese hamster cell line mutant EM9, which has a reduced ability to repair DNA strand breaks, is noted for its highly elevated frequency of sister chromatid exchange, a property shared with cells from individuals with Bloom's syndrome. The defect in EM9 cells was corrected by fusion hybridization with normal human fibroblasts and by transfection with DNA from hybrid cells. The transformants showed normalization of sister chromatid exchange frequency but incomplete correction of the repair defect in terms of chromosomal aberrations produced by 5-bromo-2'-deoxyuridine.

In vivo cytogenetic effects of the cooked-food-related mutagens Trp-P-2 and IQ in mouse bone marrow.

Sister-chromatid exchange and chromosomal aberrations were measured in vivo in mouse bone marrow following intraperitoneal injection of the cooked food mutagens, Trp-P-2 and IQ. Trp-P-2 produced a significant positive dose response for both endpoints while IQ produced only a weak but significant sister-chromatid exchange response. The relative potency of these two chemicals is similar to that seen in mammalian cells in vitro but opposite to their potency in Salmonella.

In vivo exposure to plant flavonols. Influence on frequencies of micronuclei in mouse erythrocytes and sister-chromatid exchange in rabbit lymphocytes.

No consistent increases in the micronucleus frequency were observed in bone marrow or peripheral blood erythrocytes from mice treated with quercetin, rhamnetin, neohesperidin dihydrochalcone, or hesperetin dihydrochalcone under various exposure and sampling conditions. Over the dose range of 100-1000 mg/kg, quercetin failed to increase significantly erythrocyte micronucleus frequencies either (1) in bone marrow of male mice at 6 h after the second of 2 i.p.

The relationship between sister-chromatid exchange and perturbations in DNA replication in mutant EM9 and normal CHO cells.

The majority of the high (12-fold elevated) baseline sister-chromatid exchanges (SCEs) that occur in the CHO mutant line EM9 appear to be a consequence of incorporated BrdUrd, and they arise during replication of DNA containing BrdUrd in a template strand. In normal CHO cells the alkaline elution patterns of DNA newly replicated on a BrdUrd-containing template are significantly altered compared with those seen during the replication on an unsubstituted template. The nascent DNA synthesized on such an altered template is delayed in reaching mature size, possibly because replication forks are temporarily blocked at sites occurring randomly along the template.

A CHO-cell strain having hypersensitivity to mutagens, a defect in DNA strand-break repair, and an extraordinary baseline frequency of sister-chromatid exchange.

A mutant of CHO cells (strain EM9) previously isolated on the basis of hypersensitivity to killing by ethyl methanesulfonate (EMS) is approx. 10-fold more sensitive than the parental line, AA8, to killing by both EMS and MMS. It is also hypersensitive to killing by other alkylating agents (ethyl nitrosourea and N-methyl-N'-nitro-N-nitrosoguanidine), X-rays, and ultraviolet radiation.

Mutagenic and toxic activity of environmental effluents from underground coal gasification experiments.

Using bacterial bioassays, we have screened for the presence of mutagens and toxins in extracts from groundwater, and in tar from product gas, at the sites of two Lawrence Livermore National Laboratory (LLNL) in situ experiments: Hoe Creek II and Hoe Creek III. The sites exhibited different potential biological hazards, suggesting that different gasification processes may represent different human health concerns. We found that mutagens are present in groundwater, persist for at least 2 yr after gasification has been terminated, and show a change in activity with time-possibly in parallel with changes in chemical composition.

Variation in the baseline sister chromatid exchange frequency in human lymphocytes.

The utility of the sister chromatid exchange (SCE) assay for human population studies is potentially limited by the variability associated with individual baseline SCE Frequencies. This investigation identifies and quantifies the major sources of preparative and biological variation associated with the determination of baseline SCE frequencies in cultured human lymphocytes. Much of the variation in lymphocyte SCE frequencies is attributable to the amount of bromodeoxyuridine (BrdUrd) available per lymphocyte; the pooled coefficient of variation (CV) over the dose range of 10 to 160 micrometer is about 18%.

DNA crosslinking, sister-chromatid exchange and specific-locus mutations.

Chinese hamster ovary cells were treated with the DNA-crosslinking chemicals, mitomycin C (MMC) and porfiromycin (POR), and their monofunctional derivative decarbamoyl mitomycin C (DCMMC). After exposure, the cells were studied for the induction of sister-chromatid exchanges (SCEs) and mutations at the hypoxanthine phosphoribosyltransferase and adenine phosphoribosyltransferase loci. The frequency of SCEs varied significantly in successive sampling intervals, requiring the weighting of each interval by the percentage of second-division mitosis in that interval to obtain the mean SCE frequency for each dose.

Induction of long-lived chromosome damage, as manifested by sister-chromatid exchange, in lymphocytes of animals exposed to mitomycin-C.

The cytogenetic effects of repeated vs. acute exposure to a chemical mutagen--carcinogen were determined with an in vivo system in which chemicals injected into rabbits induce sister-chromatid exchanges (SCEs). SCE induction can be monitored when the animal's peripheral lymphocytes are cultured in the presence of bromodeoxyuridine (BrdUrd) and then scored for SCE frequency.

Purification of the chromosomes of the Indian muntjac by flow sorting.

Metaphase chromosomes were isolated from a male Indian muntjac cell line, were stained with ethidium bromide and were analyzed by flow microfluorometry to establish a deoxyribonucleic acid (DNA)-based karyotype. Five major peaks were evident on the chromosomal DNA distribution corresponding to the five chromosome types in this species. The amount of DNA in each chromosome was confirmed by cytophotometric measurements of intact metaphase spreads.