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Biomedical subjects

J L Beck

Publications and source records attributed to J L Beck.

At least 37 records · Page 2Linked to original sources

Role of sequence amplification in the generation of nematode mitochondrial DNA polymorphism.

Romanomermis culicivorax, an obligate parasitic nematode of mosquitos, possesses an unusually large mitochondrial genome. Individuals are monomorphic for one of several mitochondrial DNA (mtDNA) size variants ranging from 26-32 kb. In this report, we demonstrate that the mitochondrial genome size differential in three isofemale lineages is due to the presence of mtDNA sequences amplified to different copy numbers within each mtDNA molecule. Restriction enzyme analysis and DNA sequencing studies reveal that each mitochondrial genome contains one of two 3.0 kb repeat types that differ by approximately 30 bp. This difference is primarily due to a short (23 bp) imperfect tandem duplication present within the larger of two polymorphic repeating units. The 3.0 kb reiterated DNA sequences are present as direct, tandem repeats and as inverted portions of the same sequence located elsewhere in the genome. Based on mtDNA analysis of an independently reared R. culicivorax culture, we conclude that events resulting in mitochondrial genome rearrangement occurred in natural field populations prior to propagation within the laboratory.

Animals↗

In situ hybridization as a tool to study numerical chromosome aberrations in solid bladder tumors.

Methods for single- and double-target in situ hybridization (ISH) to, cells isolated from solid transitional cell carcinomas (TCC's) of the urinary bladder are described. Single cell suspensions were prepared from solid tumors of the urinary bladder by mechanical disaggregation and fixed in 70% ethanol. Using two DNA probes specific for the centromeres of chromosomes #1 and #18, ISH procedures were optimized for these samples. Human lymphocytes and cells from the T24 bladder tumor cell line were used as controls. In lymphocyte nuclei and metaphase chromosome spreads, ISH showed two major spots for each of the probes. About 80% of the nuclei from T24 cells showed three spots for both the chromosome #1 and #18 specific probes. When nuclei from TCC's were analyzed, often the number of spots for chromosome #1, and to a lesser extent for chromosome #18, differed from the number expected on basis of flow cytometric ploidy measurements. The double target-ISH method in all cases allowed the correlation of numerical aberrations for chromosomes #1 and #18 in one and the same cell. By such analyses a profound heterogeneity in chromosome number was detected in most tumors. In order to optimize the reproducibility of the method and the interpretation of the ISH-signals, criteria for their analysis have been determined. This procedure can now be applied on a routine basis to solid tumor specimens.

Carcinoma↗

Sequence amplification and gene rearrangement in parasitic nematode mitochondrial DNA.

The nematode Romanomermis culicivorax, an obligate mosquito parasite, possesses a 26 kilobase (kb) mitochondrial genome. The unusually large size is due to transcriptionally active DNA sequences present as 3.0 kb direct tandem repeats and as inverted portions of the repeating unit located elsewhere in the mitochondrial DNA (mtDNA). The genome rearrangements involved in establishing this unusual sequence organization may have dramatically altered conventional mitochondrial gene order. Genes for subunits of the cytochrome c oxidase complex (COI and COII) are normally closely linked in animal mtDNAs, but are separated by approximately 8 kb in this mitochondrial genome.

Animals↗

Tissue-specific markers in flow cytometry of urological cancers. III. Comparing chromosomal and flow cytometric DNA analysis of bladder tumors.

Thirty-seven transitional-cell carcinomas (TCC) of the urinary bladder were analyzed by DNA flow cytometry (FCM). After labelling of the cell suspensions with antibodies to cytokeratin, the cytokeratin-positive cells and the non-epithelial cytokeratin-negative cells could be analyzed separately. After estimation of S- and G2M phase, 3/17 cases (18%) with a normal DNA index showed elevated proliferative levels, among cytokeratin-labelled suspensions only. Of these 17 cases, 14 showed chromosomal abnormalities. The remaining 20 cases were abnormal, irrespective of the technique used. Although immuno-labeling of tumor cells for cytokeratin in FCM increases the sensitivity of this method in detecting aneuploid tumors or tumors with high proliferation fractions, the discriminating power of chromosomal analysis of TCC is greater than FCM.

Carcinoma, Transitional Cell↗

Survival of spermatogonial stem cells in the rat after split dose irradiation during LH-RH analogue treatment.

A rat model has been created in which a single injection of an LH-RH analogue depot preparation (Zoladex, ICI 118630) produced a temporary interruption of the pituitary-gonadal axis. This effect applied during irradiation was investigated as a possible mechanism to protect the testis from radiation damage. A local testicular irradiation dose of 6.0 Gy was given either as a single dose or as a fractionated (2 X 3.0 Gy) dose at different time intervals ranging from 8 to 72 h. Stem cell survival was measured 11 weeks after irradiation by means of the repopulation index and the number of haploid cells (spermatids) measured by flow cytometry. Serum gonadotrophins and testosterone concentrations were measured to evaluate hormonal recovery. No significant differences were observed between serum concentrations of follicle-stimulating hormone (FSH), luteinizing hormone (LH) and testosterone and the duration of the fractionation interval. Stem cell survival was higher following fractionated irradiation in comparison with the single dose. For the 8 h interval an increase in recovery ratio was found, amounting to a factor of 5 of the single dose value. The fluctuating pattern of the recovery curves indicated changes in radiosensitivity of stem cells. The combination of hormonal inhibition of spermatogenesis and fractionated irradiation led to a decrease in the absolute numbers of stem cells. However, the stem cell recovery curves were identical to those seen without hormonal inhibition. It was concluded that hormonal pretreatment with Zoladex during split dose irradiation had no protective effect on stem cell survival.

Animals↗

A computer model of spermatogenesis in the rat; correlation with flow cytometric data based on autoradiographic cell-cycle properties.

A computer model of rat spermatogenesis was created, based on autoradiographic studies of durations of the phases of the cell cycle (G1, S, G2 and mitotic phases) of each germ-cell type. With this model it is possible to predict and to gain insight into the changes of the DNA content occurring during the normal process of spermatogenesis. The relative proportions of haploid, diploid, S phase and tetraploid germ cells with increasing age of the rats were calculated. Calculated and actual experimental flow cytometry data were compared to test the accuracy of the model, and these show good agreement. The present work demonstrates that single-parameter DNA analysis of testicular cells is primarily a reflection of germ cells in the spermatocyte and spermatid stages of development, and of non-germ cells. The FCM single-parameter DNA analysis of testicular cells is relatively insensitive to changes in the stem cell and spermatogonial stages of germ-cell development.

Animals↗

Effects of danazol on spermatogenesis in adult rats.

Adult male Wistar rats were treated with Danazol (4 mg/day s.c.) for 52 days. The drug produced a marked, rapid drop in serum testosterone concentrations to very low levels and caused a slower decrease in serum FSH, LH and testis weight. Flow cytometric analysis of testicular cell suspensions showed a decline in the absolute numbers of haploid cells (spermatids), tetraploid cells (mainly pachytene spermatocytes) and of cells in the S-phase of the division cycle, suggesting that Danazol inhibited proliferation of spermatogonia and/or primary spermatocytes. Histological counting of the different types of spermatogonia, however, revealed no significant change in their numbers during Danazol treatment. It is concluded that Danazol inhibited spermatogenesis primarily after the preleptotene stage of primary spermatocytes.

Animals↗

Effects of a single injection of a new depot formulation of an LH-releasing hormone agonist on spermatogenesis in adult rats.

Adult male Wistar rats were treated with a single injection (500 micrograms s.c.) of a new biodegradable depot formulation of the LH-releasing hormone (LHRH) analogue [D-Ser(But)6]AzGly10-LH-RH (Zoladex; ICI 118,630) to evaluate its potential for inhibiting spermatogenesis. The drug produced a marked (P less than or equal to 0.05) decrease in serum concentrations of FSH, LH and testosterone with a maximum effect 14 days after treatment. Since striking focal histological changes were seen in the testis after only 1 week, at a time when changes in serum gonadotrophins were minimal, there may be a direct effect of the LHRH analogue on spermatogenesis. Degenerative changes in germ cells as well as Sertoli cells could be observed. Flow-cytometric analysis of testicular cell suspensions showed a significant decline in the absolute numbers of haploid cells (spermatids), tetraploid cells (mainly pachytene spermatocytes) and of the numbers of cells in the S-phase of the cell cycle. This suggests that the drug also inhibits proliferation of spermatogonia and/or primary spermatocytes. Testis weight, serum hormone concentrations, and histological and cytological parameters returned to essentially normal values 52 days after the injection. It is concluded that this new method of administration may have practical and pharmacokinetic advantages for the purpose of reversible inhibition of spermatogenesis.

Animals↗

Cell DNA content--correlation with clonogenicity in the human tumour cloning system (HTCS).

Thirty-six ovarian and renal-cell tumours were analyzed by flow cytometry (FCM) for DNA content and in parallel were assayed for colony formation in a human tumour cloning system (HTCS). While 15/19 (79%) tumours with an abnormal (aneuploid) DNA stemline formed colonies in the HTCS, only 2/17 (12%) of diploid tumours formed colonies. All samples contained tumour cells as assessed by routine cytological examination. The capacity to form colonies in the HTCS was not correlated in these tumours with grade or stage of disease or tumour type. The level of aneuploidy expressed as the FCM DNA index did not correlate with the cloning efficiency in HTCS. These findings suggest that tumour growth in the HTCS reflects a biologically important potential, related in at least some tumours to an abnormal DNA stemline.

Carcinoma, Renal Cell↗

DNA analysis and sorting of rat testis cells using two-parameter flow cytometry.

By use of two-parameter flow cytometry of rat testis cell suspensions stained with mithramycin for DNA (the peak amplitude of the fluorescence signal versus total fluorescence intensity integrated over time), eight cell compartments could be distinguished without pre-enrichment of the samples. Cells in these compartments were identified by sorting and subsequent microscopic examination.

Animals↗

The female athlete's knee.

Available data have established that the female athlete is particularly vulnerable to various stress syndromes and noncontact ligament sprains of the knee. Although anatomic differences such as decreased muscle mass and increased joint laxity exist, the primary reason for this disparity appears to be an inadequate level of basic motor skills. The average female athlete has less training and conditioning than the male and has paid less attention to preparticipation fitness. Normal motor function has been defined in terms of required flexibility and coordination so that simple clinical testing can be performed. Normal flexibility may be defined as the unimpeded range of motion at each joint required for running. Coordination is normal when posture and movement are efficient and self-protective. This results from the action of primary neuromuscular mechanisms, which are anthropologic developments to support the unique posture of humans. Poor motor training or reflex inhibition causes secondary, less efficient mechanisms to be substituted for the primary ones, resulting in faulty coordination. These changes can be detected on clinical examination. Assessment of flexibility and coordination are the most important parts of the evaluation of the female athlete, since they relate directly to her most common injury patterns. The underlying neuromuscular physiology governing postural control and movement needs to be considered in planning therapeutic measures.

Athletic Injuries↗

Overuse injuries.

Stress injuries common to running athletes are always associated with a breakdown of the protective mechanisms normally inherent in healthy posture and gait. It is more important to identify and treat these breakdowns of the motor process than the resulting injury itself. Simple motor screening of athletes can identify functional abnormalities that lead to stress injuries. This screen includes assessment of flexibility of all major joints, responsiveness of primary postural muscles, posture, balance, and gait. Present definitions of physical fitness are focused on cardiorespiratory function and strength and do not properly determine the ability of an athlete to run or walk. The basic motor screen outlined in this article has been shown to be of value in correcting this shortcoming.

Athletic Injuries↗

Enzymatically active zinc, copper and mercury derivatives of the one-iron form of pig allantoic fluid acid phosphatase.

Derivatives of the violet, iron-containing acid phosphatase of pig allantoic fluid have been prepared in which one of the two iron atoms present in the native enzyme has been replaced by zinc, copper or mercury. The derivatives so formed are enzymatically active: the Zn-Fe, Cu-Fe and Hg-Fe enzymes have specific activities of about 80%, 25% and 17% respectively, of the maximum specific activity of the Fe-Fe enzyme in the standard assay at pH 4.9 with p-nitrophenyl phosphate as substrate. In contrast to the Fe-Fe enzyme, the mixed metal derivatives are not rapidly inactivated by H2O2. Visible absorption spectra of the derivatives confirm that all of the visible absorption of the Fe-Fe enzyme is due to one of the iron atoms. Attempts to prepare an active Cu-Cu enzyme were unsuccessful.

Acid Phosphatase↗