Embryonic origin of skeletal muscle cells in the iris of the duck and quail

Summary The origin of skeletal muscle cells in avian iris muscle was investigated by quantitative analysis of heterochromatin profiles at the electron-microscopic level in irides of six types of quail-duck chimeras. Each of the following tissues was transplanted into the head region from quail to duck between stages 9 and 10: cranial neural crest; trunk neural crest; midbrain and adjacent mesoderm; forebrain; forebrain without neural crest; and forebrain without neural crest and mesoderm. The average ratio of heterochromatin profile to nucleus profile in iris skeletal muscle cells was high (quail type) in the dorsal iris, but low (duck type) in the ventral iris of the chimeras resulting from isotopic transplantation of cranial neural crest. Heterotopic transplantation of trunk neural crest to cranial position resulted in failure of development of skeletal muscle cells in the dorsal iris, but not in the appearance of skeletal muscle cells in the ventral iris. The average ratio of heterochromatin profile to nucleus profile in iris skeletal muscle cells was high in the chimeras resulting from transplantation of midbrain region and the chimeras resulting from transplantation of forebrain region, intermediate in the chimeras resulting from transplantation of forebrain region without neural crest, and low in the chimeras resulting from transplantation of forebrain region without neural crest and mesoderm. These results indicate that the skeletal muscle cells in the dorsal iris are of cranial neural crest origin while those in the ventral iris are not, and could possibly arise from cranial mesoderm.     

Radioiodinated surface proteins of rabbit trophectoderm cells

Summary We have previously used surface iodination to discriminate between the protein patterns of epithelial cell surfaces in uteri of rabbits receptive (Day 6.5) or nonreceptive (Day 4) to nidation (Ricketts et al. 1984). In this paper, we describe application of the same technique to the trophoblastic surface of rabbit blastocysts collected on the same days of pregnancy. Analysis of labelled proteins by polyacrylamide-gel electrophoresis under denaturing conditions did not reveal qualitative differences between the two days of pregnancy. Scanning densitometry was used to quantitate the area under each protein peak on an autoradiogram; these areas were used as variables in statistical analysis of the protein pattern of individual animals. Quantitative differences between the protein patterns of the two surfaces were detected by canonical variate analysis of the pattern of relative areas of labelled protein peaks. In proteins separated on 7.5% gels, this statistical analysis correctly assigned blastocysts from 8 out of 10 animals to one of two groups according to day of pregnancy. The discrimination was not statistically significant, however, in protein patterns on 12.5% gels, used to give better separation in the lower range of molecular weights. The same analysis in the uterus unequivocally separated the surface iodination patterns from these same days of pregnancy. Thus the changes detected by surface iodination appear to be less pronounced on the trophectoderm than on the uterine epithelium in relation to the time of ovoimplantation.     

Age and growth-related changes in cyclopiazonic acid-potentiated lipophilic cation accumulation by cultured cells and binding to freeze-thaw lysed cells

In a previous study (1) we demonstrated that increased tetraphenylphosphonium (TPP) uptake by renal epithelial cells (LLC-PK₁) exposed to the fungal metabolite cyclopiazonic acid (CPA) was not a result of hyperpolarization across the plasma membrane even though CPA-potentiated TPP uptake could be totally inhibited by the depolarizing agent carbonylcyanide-m-chlorophenylhydrazone (CCCP). We now demonstrate that CPA potentiates TPP accumulation by proliferating skeletal muscle (L6) and LLC-PK₁ cells but not by nonproliferating primary rat hepatocytes. In LLC-PK₁ cells, CPA-potentiated TPP accumulation is observed in cells at all ages. In s cells, CPA-potentiated TPP accumulation is maximal soon after subculturing, and as the cells age they become less sensitive to CPA until TPP accumulation by CPA-treated cells approaches that of untreated cells. The temporal change in sensitivity of L6 cells to CPA may be related to biochemical and/or metabolic changes which occur as the cells age in culture. Hepatocytes, LLC-PK₁ cells, and L6 cells permeabilized by freeze-thaw lysis, all exhibit CPA-potentiated TPP partitioning, even in the presence of CCCP. This result indicates that both TPP and CPA must have access to the intracellular space in order for potentiated TPP partitioning to be observed. We hypothesize that the site of interaction between CPA and TPP is intracellular and probably associated with the cytoplasmic side of the plasma membrane and possibly the mitochondria.     

Radio frequency radiation causes no nonthermal damage in enzymes and living cells

The ability of radio frequency radiation (RFR) to exert irreversible nonthermal (i.e., not caused by accompanying heat) effects on biologics has been widely debated due to a relative paucity of comprehensive critical details in published reports dealing with this issue. In this study, we used rigorous control over experimental conditions to determine whether continuous RFR nonthermally affects commercially important enzymes and live bacterial and human cells using three most commonly used frequencies in current RF identification technology, namely 2.45 GHz, 915 MHz, and 13.56 MHz. Diverse biological samples were exposed to RFR under deliberately harsh conditions to increase the likelihood of observing such effects should they exist. Enzymatic activities of horseradish peroxidase and β‐galactosidase in aqueous solution exhibited no statistically discernable consequences of even very intense RFR. Likewise, with putative thermal effects excluded, the viabilities of bacteria (both gram‐positive and gram‐negative) and of human cells were not detectably compromised by such an RFR exposure. © 2010 American Institute of Chemical Engineers Biotechnol. Prog., 2010     

siRNA干扰PDGF对新生儿血管瘤干细胞的增殖和分化的影响

婴儿血管瘤是一种血管瘤,表现出独特的快速生长的特征,然后随着时间而消退。血管瘤来自CD133+干细胞,当植入免疫缺陷小鼠时,它们分化成内皮细胞。同样克隆扩增的干细胞也产生脂肪细胞,从而重现血管瘤的消退期。本研究主要阐明了使用血管瘤来源的干细胞(hemSC)增殖和分化的内在机制。本研究发现血小板衍生生长因子(PDGF)在增殖期升高并可能抑制脂肪细胞分化。hemSC表达高水平的PDGF-b并且在基础(未刺激)条件下显示PDGF受体的持续酪氨酸磷酸化。PDGF受体信号传导的抑制导致hemSCs中的脂肪生成增强。此外,hemSCs暴露于外源性PDGF-b降低了脂肪含量和脂肪细胞特异性转录因子的表达。总之,本研究将PDGF信号传导鉴定为血管瘤退化的内在负调节因子,并强调了破坏PDGF信号传导治疗血管瘤的治疗潜力。     

Influence of long term treatment with 3-methylcholanthrene or carbaryl on mixed-function oxidase activity in 3T3 fibroblasts: Studies by microspectrofluorimetry on single cells

Using benzo(a)pyrene (BaP) as a probe for aryl hydrocarbon hydroxylase (AHH) activity, differences in mixed-function oxidase (MFO) activity were observed using microspectrofluorimetry in single living cells during long term treatment with 3-methylcholanthrene (3-MC) or carbaryl. Although these two compounds differ in chemical structure, similar effects were observed in 3T3 cell populations. The results suggest that the two compounds activate the same enzymatic system and that individual cells of a supposed homogeneous cell population are not equally sensitive to xenobiotics, i.e. subpopulations were observed which have differences in AHH activity.     

Ehrlich ascites tumour cells become refractory to alpha-difluoromethylornithine at a certain stage of growth

When Ehrlich ascites tumour cells are induced to proliferate by serum stimulation, the ornithine decarboxylase (ODC) activity increases rapidly and reaches two to three peaks during the first 24 h. Inhibition of the first peak in ODC activity (occurring at 4 h) by adding alpha-difluoromethylornithine (DFMO) within 2 h of serum stimulation, results in maximal growth inhibition. Under these conditions, similar degrees of polyamine depletion are achieved. When DFMO is added 3 h after seeding, however, enough polyamines have already accumulated during the initial burst in ODC activity to reduce the antiproliferative effect of the drug. The antiproliferative effect is further reduced when DFMO is added 6 h after seeding. When DFMO is added 23 h after seeding, i.e. after maximal accumulation of polyamines, there is no inhibition of cell proliferation. These findings are important to consider both when designing experimental as well as clinical regimens for this drug.     

Nucleotide pool changes in coelomic cells (eleocytes) of the polychaete Nereis virens during sexual maturation

Eleocytes (a type of coelomic cell) of the polychaete Nereis virens can store large amounts of adenine nucleotides at certain times. Since eleocytes have specific functions related to gametogenesis, we tested whether the presence of these large nucleotide stores in eleocytes is specific to gender or related to specific events during gametogenesis. Nucleotide pools in eleocytes isolated at different stages of sexual maturation from N. virens were analysed using high-performance liquid chromatography. Eleocytes from immature and male animals had extremely high concentrations of both AMP and ADP (each > 10 μmol/ml of packed cell volume). In eleocytes from male animals, the high nucleotide stores were maintained throughout the maturation phase and decreased at a late stage, while in female animals the nucleotides were degraded at an early stage of maturation. In male eleocytes, the decrease in the adenine nucleotide pool may be the result of its conversion to inosine which is then released by the eleocytes and reutilized by male germ cells for nucleic acid biosynthesis, as has been suggested previously. Our study shows that the time of degradation of the adenine nucleotide pool coincides with the period of spermatogonia proliferation which involves intense nucleic acid synthesis. ATP levels (0.4–1.5 μmol/ml packed cell volume) and the guanine nucleotide pool (GTP+GDP+GMP; 0.08–0.18 μmol/ml packed cell volume) were similar in both sexes, did not change during germ cell development and were decreased only in eleocytes from prespawning females. The GTP/GDP ratios were initially higher (up to 14) in eleocytes from females compared to ratios in eleocytes from immature (4–9) and male animals (up to 8), and decreased during the maturation phase of the animals. GTP levels were correlated with those of ATP; this correlation was much closer in eleocytes from females than from males. The results further support the hypothesis that the adenine nucleotide stores in the eleocytes are maintained as a supply of purine precursors for the growing germ cells.