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At least 55 records · Page 3Linked to original sources

PubMed2001

Ion-dependence of Z-line and M-line response to calcium in striated muscle fibres in rigor.

The calcium-dependent contraction of vertebrate skeletal muscle is thought to be primarily controlled through the interaction of the thick and thin filaments. Through measurement of the Donnan potential, we have shown that an electrical switching mechanism (sensitive to both anions and cations) is present in both A- and I-bands [1]. Here we show that this mechanism is not confined to the contractile apparatus and report for the first time the presence of M-line potentials. The Z-line responds to Ca2+ ions in a similar manner to the A-band under the same solution conditions (phosphate-chloride and imidazole buffers), even though it has no reported Ca2+ binding sites. Z-line potentials were not observed in tris-acetate buffer. The M-line has a markedly different response to any of the other subsarcomeric regions, however, and can only be detected in the phosphate-chloride buffer. Preliminary observations of the M-line potential in creatine kinase-deficient mouse muscle (phosphate-chloride buffer) reveal significant differences in the calcium-induced transitions between two of the genotypes and demonstrate definitively that it is the M-line potential that is being recorded. From these results, it seems likely that the charge response of the Z-line and M-line is being mediated by titin in an anion-dependent manner. Our evidence comes from several observations. First, the similarity between the response of the Z-line potentials to the A-band potentials, where titin is the only link between these structures and second, the differential observation of M-line and Z-line potentials in a range of buffers containing different anion(s). Both Z-line and M-line potentials were seen in phosphate-chloride buffer, but only the Z-line potentials could be detected in chloride-only (imidazole) buffer and neither was observed in the acetate buffer. The latter observations can be attributed to two sources. The first is the effect of acetate buffer on the conformation of myosin [2]; the second is the absence of binding of the M-line protein, myomesin, to titin in the absence of phosphate ions [3].

Acetates↗

PubMed1997

Genetics of growth and reproduction in the turkey. 13. Effects of repeated backcrossing of an egg line to two sire lines.

The objective of the present study was to determine the feasibility of rapidly increasing the BW of dam lines by repeated backcrossing of a dam line to a sire line. A line (E) selected long-term for increased egg production was backcrossed to a line (F) selected long-term for increased 16-wk BW (BC1) and to a commercial sire line (BC2). The BW of the F and commercial sire lines were about twice as large as that of the E line. Egg production of the E line was more than double the egg production of either of the sire lines. Large gains in BW and breast width were observed in both backcrosses. Based on additive genetic variation, gains in BW were less than expected in the first generation for females in the BC1 and for both sexes in the BC2. Values observed for males of the BC1 were similar to expected values at 8 and 16 wk and larger than expected values at 20 wk of age. Slower than expected BW gains in Generation 1 may have been attributable to the small egg size of E females. After Generation 1, BW at 16 and 20 wk of age were equal to, or greater than, expected. Egg production was reduced in both backcrosses relative to the E line. Fertility and hatchability were reduced relative to the E line by backcrossing in BC1 but not in BC2. Results of the present study suggest limited repeated backcrossing of a dam line to a sire line may be an economically feasible method to greatly increase the BW of dam lines without unduly sacrificing reproduction capability.

Aging↗

PubMed2001

Evaluation of between-line variation for within-line selection against ascites in broilers.

Within-line opportunities for selection against ascites were studied in a data set comprising a 10-line comparison. The study attempted to reveal whether contrasts between lines provide reliable candidate traits for within-line selection. Mortality was chosen as the reference trait. As no pedigree information was available, a trait was required that related mortality to the nonmortality data. By principal component analysis, such a trait (ASC_INDIC = ascites indicator) was developed from pathology data. The composite trait ASC_INDIC ranked lines well for their mortality figures (r = 0.96), from which it was concluded that ASC_INDIC represents an underlying continuous ascites trait. Between lines, blood gas traits seemed to be the most promising traits. Within lines, performance traits appeared to be highly correlated to ascites. Comparison of within-line variation to between-line contrasts revealed considerable differences. The high correlation of the blood gas traits with mortality was not present within lines. However, although the magnitude was considerably reduced, the nature of the blood gas traits in their relationship to ascites was similar within and between lines. The study primarily demonstrates that contrasts between lines carry systematic but limited information for within-line coherence. Therefore, line contrasts must be interpreted with care when aiming to study genetic variation and coherence within lines.

Animal Husbandry↗

PubMedDate not supplied

Genetic similarity and relationships of DNA fingerprints with performance and with heterosis in Japanese quail lines from two origins and under reciprocal recurrent or within-line selection for early egg production.

DNA fingerprints of Japanese quail male and female pure line breeders were obtained with probes 33.6, 33.15, and R18.1 and they yielded a total of 59 scoreable bands. Bandsharing (0<BS<1) was calculated within and between six quail lines of two origins, and under reciprocal recurrent (AA and BB), within-line (DD and EE) or no (PP and FF) selection. Twenty one pair types were compared. BS was 0.30 higher within line than between lines. BS with the control line was smaller for reciprocal recurrent selection lines than for lines under individual selection. Bandsharing between the two reciprocal recurrent selection lines was 0.19 lower than between lines under individual selection. These results indicate that the two selection methods had different effects on the genetic constitution of the lines, in agreement with previous observations made from the analysis of biochemical polymorphisms with the same set of birds. Egg production and weight traits of pure and crossbred progeny from fingerprinted quail were obtained and compared, and a linear relationship with the measure of bandsharing was estimated. No significant regression coefficient of performance on BS was found over all progeny genetic types. Heterosis from individual matings could also be estimated under the two selection methods since the same birds were parents of both pure and crossbred performance-tested quail. The association of heterosis with the difference between BS of parents of the purebreds and BS of parents of their half-sib crossbreds was favourable and significant for early production traits in lines DD and EE, but no relationship was found in lines AA and BB. These results indicate that the high level of heterosis obtained through reciprocal recurrent selection, and the heterosis observed under within-line selection may have, partly at least, a different genetic determinism. Therefore, the relationship of heterosis with BS may also depend on the past history of selection in the lines.

Journal Article↗

PubMed1983

Use of fluoresceinated complement-coated bacteria and sheep erythrocyte-antibody-complement complexes for identification of complement receptors on lymphoid cell lines: differences in binding characteristics between cell lines of normal and malignant origin.

Twenty-four lymphoma-derived cell lines, 11 cord blood lymphocyte lines, and 3 lymphoblastoid cell lines derived from normal adults were examined for complement (C) receptors utilizing fluoresceinated C-coated bacteria (FBC) to determine the optimal conditions for each type of cell line. Incubation of FBC with lymphoma-derived cell lines at 37 and 0.5 degrees C showed that maximal FBC binding at both temperatures was after 120 minutes, and peak reactivity was invariably higher at 37 degrees C. These temperature-dependent differences were similar, both in Epstein-Barr virus nuclear antigen (EBNA)-positive and EBNA-negative lines. EBNA-positive lines, however, expressed higher levels of FBC rosettes than EBNA-negative lines at both temperatures. In contrast, FBC binding to cord blood cell lines after 120-minute incubation was maximal at 0.5 degrees C. Although similar numbers of FBC rosettes were formed after 30 minutes at both 37 and 0.5 degrees C in cord blood cell lines, rosette formation deteriorated after longer periods of incubation at 37 degrees C. The optimal temperature for FBC binding to lymphoblastoid cell lines could not be determined, since bacteria bound spontaneously to these lines at 37 degrees C. Cell lines were also tested simultaneously for sheep erythrocyte-antibody-complement complex (EAC)M and FBC binding at 37 and 0.5 degrees C. FBC reactivity under optimal conditions for each type of cell line correlated well with EACM reactivity at 37 degrees C. The significance of these results is discussed.

Burkitt Lymphoma↗

PubMed1984

Quantitation of fibrinogen influx and fibrin deposition and turnover in line 1 and line 10 guinea pig carcinomas.

Radiolabeled guinea pig fibrinogen (GPF) was used to measure fibrinogen influx and fibrin accumulation in line 1 and line 10 hepato- (bile duct) carcinomas growing in the s.c. space of syngeneic strain 2 guinea pigs over the course of 7 days following transplant, an interval of growth uncomplicated by immunological tumor rejection or by significant tumor necrosis. Earlier immunofluorescence studies revealed fibrin deposits in both tumors with line 1 much greater than line 10. In accord with these data, GPF accumulated in both tumors in amounts that matched or exceeded plasma fibrinogen levels. Line 1 tumor GPF content was 4-fold greater than that of line 10 tumors and 11- to 33-fold that of normal s.c. tissue. The composition of tumor fibrinogen-fibrin was investigated by aqueous and urea extraction. The fraction of total accumulated GPF that was urea insoluble, and therefore presumably cross-linked fibrin, was constant over time but strikingly different for line 1 (65%) and line 10 (48%) tumors, as compared with control s.c. tissue (18%). By 7 days, line 1 tumors (mean weight, 0.77 g) contained nearly 2 mg of fibrinogen-fibrin, and line 10 tumors (mean weight, 0.62 g) contained nearly 0.5 mg. Influx of GPF and initial clotting were constant over time and equivalent for the two tumors. Hence, the large differences in GPF accumulation observed between these tumors apparently reflect differences in fibrinolysis, not in fibrinogen influx or coagulation. The data presented indicate substantial traffic of plasma fibrinogen into and out of both tumors, as compared with control tissues, equivalent to nearly 10 and 7 ml of plasma over 7 days of growth for line 1 and line 10 tumors, respectively; comparable values for normal s.c. tissues were 1.0 to 1.4 ml plasma fibrinogen. Even in line 1 tumors with their abundant fibrin gel, only 6.3% of GPF entering tumors over 7 days was retained, as compared with 2% for line 10 tumors and approximately 1% for control tissue.

Animals↗

PubMed1994

Functional insulin and insulin-like growth factor-1 receptors are preferentially expressed in multiple myeloma cell lines as compared to B-lymphoblastoid cell lines.

While IGF-1 plays a role in early B-cell development, little is known of insulin and insulin-like growth factor-1 (IGF-1) action in post-marrow B-cells. Recently, our laboratory demonstrated that mouse and human multiple myeloma (MM) cell lines possess functional insulin receptors (IRs) and IGF-1 receptors (IGF-1Rs). In this study, we show that responsiveness to insulin and IGF-1 is more developed in human MM cell lines than in human B-lymphoblastoid cell lines. Two human MM cell lines (U266 and RPMI 8226) were compared to three B-lymphoblastoid cell lines [Epstein-Barr virus immortalized B-cells (EBV), a Burkitt lymphoma cell line (Ramos), and a non-EBV lymphoblastoid cell line (HS Sultan)]. Surface IR and IGF-1R expression, measured by flow cytometry, demonstrated that the MM cell lines expressed more IRs and IGF-1Rs than did the EBV, Ramos, or HS Sultan cell lines. In vitro receptor kinase activity of affinity-purified receptors showed that the MM cells had more phosphorylated receptors than did the EBV, Ramos, or HS Sultan cells. Intracellular receptor signaling was also markedly different between the two cell groups. Whole cell phosphorylation studies showed that MM cells possessed not only hormone-dependent receptor autophosphorylation (M(r) 97,000) but also substrate phosphorylation (M(r) 185,000; 60,000). The lymphoblastoid cells, while demonstrating receptor autophosphorylation (IR autophosphorylation in the EBV cell line at 200 nM hormone was similar to MM receptor phosphorylation at 2 nM), lacked hormone-responsive substrates. The MM cell lines contained significantly more hormone-stimulated phosphatidylinositol 3-kinase (PI 3-kinase) activity than the B-lymphoblastoid cell lines. In the MM cells, PI 3-kinase was activated by at least 10-fold, but, in the B-lymphoblastoid cell lines, it was activated by no more than 2-fold. Hormone-responsive glucose metabolism was also greater in the MM cell lines. In the U266 cells, insulin increased lactate production 62 +/- 9 and 101 +/- 12% (mean +/- SE) at concentrations of 2 nM and 200 nM, respectively. IGF-1 produced 72 +/- 9 and 99 +/- 13% increases at similar concentrations. In the 8226 cells, insulin increased lactate production 4 +/- 4 and 36 +/- 15% at 2 and 200 nM, respectively. IGF-1 produced a 13 +/- 6 and 70 +/- 18% increase. In the EBV and Ramos cells, neither hormone increased lactate production by more than 10 +/- 3%.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗

PubMed2005

Crosstalk between on-line and off-line processing of visual features.

Three dual-task experiments were conducted to investigate the relation between immediate, "on-line" judgments about visual features and delayed, "off-line" judgments. One hypothesis ("selective interference") predicted that dual-task performance would be challenged specifically within a visual dimension, as both tasks compete for the same resources. Another hypothesis ("cost of switching") made the opposite prediction. In Experiment 1, participants performed either color or shape discriminations in the on-line and off-line visual tasks, with systematic variation of feature similarity between the on-line and off-line features. In Experiment 2, participants performed either color or shape discriminations in the off-line task and color discriminations in the on-line task, with no overlap between the on-line and off-line features. In Experiment 3, participants performed color discriminations in both the on-line and off-line tasks, with partially overlapping stimulus sets. Altogether, the data from the three experiments provided evidence in favor of the hypothesis of cost of switching. Stimulus-stimulus compatibility effects between features in the off-line task and those in the on-line task further underscored the perceptual nature of the crosstalk.

Discrimination, Psychological↗

PubMed1985

Fine structure of wide and narrow vertebrate muscle Z-lines. A proposed model and computer simulation of Z-line architecture.

A model of the structure of vertebrate Z-lines and Z-line analogs is introduced and supported by evidence from electron microscope studies of wide Z-lines (rat and feline soleus, and feline and canine cardiac muscles), narrow Z-lines (guppy, newt and frog skeletal muscles), and Z-rods (from a patient with nemaline myopathy and from cardiac muscles of aged dog). The model is based on a pair of Z-filaments (termed a Z-unit), which are linked near their centers at a 90 degrees angle and form bridges between neighboring antipolar thin (actin) filaments. A square lattice of four Z-filament pairs (the basic structure of the Z-line, termed a Z-line unit) defines the geometrical position of the I-square unit. In this native state of the Z-line, small square and large square net forms appear in cross-section. Other cross-sectional patterns of Z-lines, including basket-weave and diagonal-square net patterns, can be explained by detachment of the Z-filament from the Z-filament binding region within each Z-filament pair due to chemical or physical stress. Dissection of Z-lines and Z-line analogs with calcium-activated neutral protease provides evidence that the width of all wide Z-line structures is determined by the amount of overlap of antipolar thin filaments from adjacent sarcomeres. Longitudinal patterns of narrow and wide Z-lines are shown and described in relation to the model. To test the proposed model, the dynamics of the Z-line unit structure were computer-simulated. An attempt was made to correlate longitudinal (z direction) and cross-sectional (x and y directions) patterns and to determine the amount of movement of thin or Z-filaments that is required to explain the diversity observed in cross-sectional patterns of Z-lines. The computer simulations demonstrated that the structural transitions among the small square, and therefore large square net, as well as basket-weave and diagonal-square net forms seen in cross-sections could be caused by movements of thin filaments less than 10 nm in any direction (x, y or z).(ABSTRACT TRUNCATED AT 400 WORDS)

Actins↗

PubMed1988

The HuT series of 'carcinogen-transformed' human fibroblast cell lines are derived from the human fibrosarcoma cell line 8387.

In 1977 Kakunaga reported the carcinogen-induced transformation of the diploid human fibroblast cell line KD into focus-forming, morphologically altered cells. Cell lines were developed from 15 individual foci. These exhibited an infinite lifespan in culture and all those that were tested (7/7) formed malignant tumors (sarcomas) in athymic mice. The existing cell lines, designated HuT-11 to HuT-14, have been studied intensively during the past decade as examples of human fibroblasts malignantly transformed by treatment with a chemical carcinogen, 4-nitroquinoline-1-oxide. Recently, in comparing the HuT-11, HuT-12 and HuT-14 cell lines with KD cells, McCormick and Maher (Mutat. Res., 199, 273-291, 1988) found evidence that the malignant cells could not have been derived from the latter. But, this did not rule out the possibility that as the target cells for his original study of carcinogen-induced transformation, Kakunaga had inadvertently used cells from some other, unidentified normal individual. Since the donor of such cells would not be known and the original cell line was not available, it would be impossible to determine the degree of identity between such a target cell line and the HuT cell lines. However, in the course of examining methods for such testing, we recently became aware that the isozyme pattern of these HuT cell lines was identical to that of the human fibrosarcoma-derived cell line 8387 established in 1966. We here report that the HuT cell lines and the 8387 cell line also exhibit an identical series of HLA determinants and identical restriction fragment length polymorphisms (RFLPs). Assuming that each of these three assays measures independently inherited characteristics, the chance that an unrelated donor of the fibroblasts that gave rise to the HuT cell lines happened to possess characteristics identical to those of the patient whose fibrosarcoma gave rise to the 8387 cell line is 1 x 10(-8). Therefore, we conclude that 8387 cells are the source of the malignant cells designated HuT from Kakunaga's original transformation experiment. Additional RFLP analysis, using a probe made from M13 bacteriophage DNA which detects a hyperpolymorphic 'minisatellite' pattern in human DNA, also showed that DNA from HuT-14 cells and from 8387 cells exhibit identical banding patterns, indicating that the cell lines were taken from the same individual. The latter banding patterns differed from that observed with DNA from KD cells.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗

PubMed2001

Ovarian steroid hormone production in a multiple ovulating male line and a single ovulating traditional line of turkeys.

Ovarian production of oestradiol and progesterone was investigated in a single ovulating traditional line and a multiple ovulating male line of turkeys. The oestradiol output from small follicles in response to LH stimulation and the aromatase activity of the residual ovary were also compared. The multiple hierarchy of follicles in the male line was shown to consist of a series of follicles of a similar size and stage of physiological maturation. The follicles of the traditional line produced significantly greater quantities of oestradiol than those of the male line. Impaired oestradiol production may have resulted in the lower plasma oestradiol concentration observed in the male line. There was no difference in follicular progesterone output between the traditional-line and male-line turkeys. The oestradiol output of whole small white follicles in response to LH stimulation was similar in the two lines and the depressed steroidogenesis of the male line was not changed. The aromatase activity of the small follicles was also lower in the male line, and there was no evidence that the male line ovary produced oestradiol in quantities proportional to its size. The results demonstrated that the hierarchy of follicles in the male line consisted of groups of follicles of similar mass and hormone output and indicated that selection for increased meat yield may have resulted in reduced ovarian steroidogenesis in male-line turkeys, in comparison with traditional, unselected turkeys.

Animals↗

PubMed1995

Nerve growth factor stimulates clonal growth of human lung cancer cell lines and a human glioblastoma cell line expressing high-affinity nerve growth factor binding sites involving tyrosine kinase signaling.

The growth of a panel of 22 different human tumor, leukemia, and lymphoma cell lines was examined in a human tumor cloning assay in agar or methylcellulose and a tritiated thymidine uptake assay. The cultures were performed in the absence or presence of increasing concentrations (0.5-500 ng/ml) of nerve growth factor (NGF). The growth of 17 of the 22 cell lines was not significantly and reproducibly affected by NGF. There was minor (1.2-fold) but reproducible stimulation of clonal growth in one glioblastoma cell line (86-HG-39) by NGF, but in this cell line NGF induced no growth modulation in a tritiated thymidine uptake assay. However, clonal growth of another glioblastoma cell line (87-HG-31) and all three lung cancer cell lines tested (HTB 119, HTB 120, CCL 185) could be stimulated up to 3-fold by NGF with a dose-response relationship for the growth factor. Growth stimulation by NGF could be completely reversed by neutralizing anti-NGF antibody and by the tyrosine kinase inhibitor genistein. Evaluation of secondary plating efficiency revealed the stimulation of colony formation as representing self-renewal and not terminal differentiation. Reverse transcriptase-PCR experiments in the five responding cell lines showed expression of both low-affinity NGF receptor (glycoprotein 75) and c-trk transcripts on the mRNA level. Of the five responding cell lines, only 86-HG-39, the cell line with the lowest responsiveness, revealed low-affinity NGF receptor on the protein level; the other four cell lines with high responsiveness, including the three lung cancer cell lines, expressed no low-affinity NGF receptor as shown by fluorescence-activated cell sorter analysis and immunoprecipitation using the ME 20.4 antibody. Immunoprecipitation using anti-trk antibodies was negative in all five responding cell lines. However, binding studies with iodinated NGF showed only low-affinity binding on the 86-HG-39 cell line and only high-affinity binding on the high-responder cell lines CCL 185 and 87-HG-31. In summary, our data suggest that NGF can be operative in stimulation of clonal growth of malignant tumor cells. High-affinity but not low-affinity binding sites mediate signal transduction for clonal growth and signaling involves tyrosine kinase activity.

Base Sequence↗

PubMed1996

Sensitivity to dexamethasone and absence of bcl-2 protein in Burkitt's lymphoma cell line (Black93) derived from a patient with acute tumor lysis syndrome: comparative study with other BL and non-BL lines.

An Epstein-Barr virus (EBV)-negative Burkitt's lymphoma (BL) cell line, designated Black93, was established in culture from a patient who developed acute tumor lysis syndrome (ATLS). Growth inhibition in vitro by dexamethasone (DXM) and the expression of bcl-2 protein (Bcl-2) were investigated in Black93 and 17 other cell lines derived from EBV-negative or -positive BL, pre-B acute lymphoblastic leukemia (ALL), follicular lymphoma (FL), and EBV-positive lymphoblastoid cell lines of normal B cell origin (B-LCL), assuming an inherent susceptibility of Black93 to cell death. The most marked growth inhibition by DXM was observed in Black93, two other BL, two pre-B-ALL and two FL lines. The other cell lines were less sensitive or were resistant. DNA extracted from the Black93 cells treated with DXM showed a ladder of oligo-nucleosomal DNA on electrophoresis. On testing of fixed smears by indirect immunofluorescence, bcl-2 protein (Bcl-2) was undetectable in Black93 and three BL lines but was detected in all the other cell lines at varying intensity. Western blot analysis showed mostly the same results. In the BL lines, the most DXM-sensitive cell lines lacked Bcl-2 expression, and the DXM-resistant cell lines always expressed Bcl-2. While none of the DXM-resistant cell lines lacked Bcl-2 expression, several pre-B or FL lines that expressed [correction of expessed] Bcl-2 were sensitive to DXM. Black93 is the first reported cell line established from a patient with ATLS. The positive sensitivity to DXM and the lack of Bcl-2 expression observed in Black93 are a major characteristic exhibited frequently by BL lines and, probably, by fresh BL cells. These properties may contribute to the precipitation of ATLS.

Acute Disease↗

PubMed1974

Establishment of a tetraploid, immunoglobulin-producing cell line from the hybridization of two human lymphocyte lines.

We here report the establishment of a seemingly permanent hybrid cell line formed by fusion of the cells of two biochemically mutant human lymphocyte lines. One parental line (UM-1-6TGr) was deficient in hypoxanthine-guanine phosphoribosyl transferase (IMP: pyrophosphate phosphoribosyltransferase, EC 2.4.2.8), and had two marker chromosomes. The second parental line (UM-21-5) was a clonal derivative of a citrullinemic lymphocyte line, and was, like the line of origin, dificient in argininosuccinic acid synthetase [(L)-Citrulline: (L)-aspartate ligase (AMP-forming), EC 6.3.4.5]. This line also had a marker chromosome, which was a B5 with a very prominent secondary constriction. After trypsinization of both parental lines, followed by addition to the fusion mixture of beta-propiolactone-inactivated Sendai virus, the cells were placed in a doubly selective medium (hypoxanthine-aminopterin-thymidine-containing medium in which the arginine was replaced with citrulline) to prevent the proliferation of the mutant parents. Under selective conditions, 97-99% of cells were found to be tetraploid, containing the three marker chromosomes; and the specific activities of the hybrid line transferase and synthetase were intermediate between normal and mutant line values. Furthermore, the UM-1-6TGr and UM-21-5 lines were producers of gamma and mu heavy chains of immunoglobulin, and of kappa light chains, as determined by immunodiffusion and immunofluorescence, and the hybrid line continued to synthesize and to secrete detectable levels of these same immunoglobulins. These studies demonstrate the genic and cytogenetic stability of this hybridized lymphocyte cell line, and prove that hybridization per se does not extinguish the activity of either the regulatory of structural genes involved in immunoglobulin synthesis.

Cell Fusion↗

PubMed2005

In vitro culture of human osteosarcoma cell lines: a comparison of functional characteristics for cell lines cultured in medium without and with fetal calf serum.

PURPOSE: Experimental in vitro models including well-characterised cell lines can be used to identify possible new therapeutic targets for the treatment of osteosarcoma. Culture media including inactivated serum is often recommended for in vitro culture of osteosarcoma cells, but the serum component then represents a nonstandardised parameter including a wide range of unidentified mediators. To improve the standardisation we have investigated whether serum-free culture media can be used in experimental in vitro studies of osteosarcoma cell lines. METHODS: The seven osteosarcoma cell lines Cal72, SJSA-1, Saos-2, SK-ES-1, U2OS, 143.98.2, and KHOS-32IH were cultured in vitro in various serum-free media and media supplemented with 10% heat-inactivated fetal calf serum (FCS). RESULTS: Although proliferation often was relatively low in serum-free media (X-vivo 10, X-vivo 15, X-vivo 20, Stem Span SFEM), some cell lines (Cal72, KHOS-32IH, Saos-2) showed proliferation comparable with the recommended FCS-containing media even when using serum-free conditions. The optimal serum-free medium then varied between cell lines. We also compared 6 different FCS-containing media (including Stem Span with 10% FCS) and the optimal FCS-containing medium varied between cell lines. However, all cell lines proliferated well in Stem Span with FCS, and this medium was regarded as optimal for four of the lines. FCS could not be replaced by fatty acids or low density lipoprotein when testing the Stem Span medium. The release of a wide range of soluble mediators showed only minor differences when using serum-free and FCS-containing media (including Stem Span with and without FCS), and serum-free Stem Span could also be used for in vitro studies of mitogen-stimulated T cell activation in the presence of accessory osteosarcoma cells. The use of Stem Span with 10% FCS allowed the release of a wide range of chemokines by osteosarcoma cell lines (Cal72, SJSA-1), and the chemokine release profile was very similar to the fibroblast lines Hs27 and HFL1. CONCLUSIONS: Serum-free culture media can be used for in vitro studies of several osteosarcoma cell lines, but the optimal medium varies between cell lines and thus depends on: (i) the cell lines to be investigated/compared; (ii) the functional characteristic that is evaluated (proliferation, cytokine release); and (iii) whether coculture experiments are included.

Acute Disease↗

PubMed1996

A prospective study of in-line skating: observational series and survey of active in-line skaters--injuries, protective equipment, and training.

OBJECTIVES: 1) To assess the relationship between types of injuries incurred and training and protective equipment worn by adults injured while in-line skating; 2) to observe the type and amount of protective equipment worn by in-line skaters while skating; and 3) to survey active in-line skaters about formal training, protective equipment, and history of injuries incurred, and the effect of such injuries on the protective equipment subsequently worn. METHODS: A prospective study of consecutive adult patients presenting to the ED for evaluation of in-line skating injuries; a consecutive-series observational study of active in-line skaters to assess protective equipment worn; and a survey of selected active in-line skaters. Eighty-five adult patients were included who presented with a history of injury related to in-line skating to the EDs of an urban academic medical center, a suburban academic-affiliated hospital, and a community hospital. Four hundred eleven active in-line skaters on the Chicago lakefront were observed for protective equipment worn, 91 of whom participated in the survey. RESULTS: Of those presenting to the ED with injuries, only 15% indicated that they had received formal in-line skating instruction. Of the ED patients, 50% wore no protective equipment; overall, 6% wore a helmet; 44%, wrist protection; 23%, knee protection; and 19%, elbow protection. Only 2% wore all of the above equipment. The primary mechanism of injury reported was a loss of balance (58%); others included collision with objects (25%), collision with bicycles (11%), and collision with cars (5%). Fractures or dislocations occurred in 48% of the patients; 6% had head injuries necessitating CT scans. Those who wore no protective gear were more likely to require hospital admission (p < 0.05). Of the 411 in-line skaters observed, 157 (38%) wore no protective equipment. Compared with the injured group presenting to the ED, fewer observed participants were without protection (p < 0.05). Among those surveyed, prior injury was not associated with the subsequent use of protective gear. CONCLUSION: Patients who present to the ED for evaluation of in-line skating injuries have a high incidence of fractures/dislocations. Few injured or surveyed in-line skaters had formal training. Use of protective equipment by injured skaters was associated with a decreased likelihood of hospitalization. Observed in-line skaters more commonly wore protective gear than did those who presented to the ED with injuries.

Athletic Injuries↗

PubMed1996

Comparison of circulating concentrations of reproductive hormones in boars of lines selected for size of testes or number of ovulations and embryonal survival to concentrations in respective control lines.

The objectives of this study were to determine whether circulating concentrations of genadotropins and gonadal hormones of boars were altered as a result of selection of pigs for size of testes or for embryonal survival and(or) number of ovulations. Included in Exp. 1 and 2 were boars with the greatest estimated paired weight of testes (TS) and boars from a control (C) line. Concentrations of FSH were similar (P > .10) in boars from the TS and C lines. In Exp. 3, 4, and 5, circulating concentrations of FSH and 17 beta-estradiol (E2) were evaluated in neonates, during pubertal development, and in mature boars of lines selected for an index of number of ovulations and embryonal survival (I), and data were compared to those for boars from a respective C line. Concentrations of E2 were not different in boars from the I line and those from the C line during the early neonatal period but were greater (P < .05) in boars of the C line than in those from the I line during pubertal development. Concentrations of FSH were greater (P < .05) in mature boars from the I line than in those from the C line. In summary, selection for size of testes did not influence circulating concentrations of FSH in mature boars. The secretory pattern of E2 in boars before puberty changed as a result of selection for embryonal survival and number of ovulations in females of the I line, and the different patterns of circulating E2 early in life may result in enhanced circulating concentrations of FSH in adult boars of the I line compared with boars of the C line.

Animals↗

PubMed2002

Indole-3-carbinol inhibits protein kinase B/Akt and induces apoptosis in the human breast tumor cell line MDA MB468 but not in the nontumorigenic HBL100 line.

We have identified a new target for the chemopreventive dietary agent indole-3-carbinol (13C) in the antiapoptotic signaling pathway involving phosphatidylinositol 3'-kinase and protein kinase B (PKB)/Akt. 13C inhibited phosphorylation and activation of PKB in the tumor-derived breast cell line MDA MB468, but not in the immortalized breast line HBL100. We propose that this cell type-specific response to 13C contributes to the differential induction of apoptosis and sensitivity to growth inhibition of the two cell lines (approximate IC50 = 30 microM for the MDA MB468 line, compared with 120 microM for the HBL100 line). 13C only induced apoptosis in the MDA MB468 cell line, but at higher doses, it increased necrosis in the HBL100 line. The tumor cell line was also markedly less able to recover when 13C was removed from the culture medium. Downstream of PKB, 13C decreased nuclear factor kappaB DNA binding, independently of an effect on IkappaB kinase, in the MDA MB468 cell line only. The tumor suppressor PTEN, which prevents phosphorylation and activation of PKB, was expressed in HBL100 cells but was not detected in MDA MB468 cells. In corroboration of the results obtained with the breast cell lines, 13C decreased phospho-PKB levels and induced apoptosis in the prostate cell line LNCaP, which expresses very low levels of PTEN, but did not do so in PTEN-positive DU145 cells. 13C did not affect PTEN levels in any cell line. This is the first study to report a differential mechanistic response of tumor-derived and nontumorigenic cell lines and of PTEN high- and low-expressing cells to 13C and indicates a promising chemopreventive role for 13C against estrogen receptor-alpha-negative, aggressive-phenotype breast tumors.

Anticarcinogenic Agents↗