Search PubMed⌕ Search

Biomedical subjects

T L Timme

Publications and source records attributed to T L Timme.

At least 55 records · Page 3Linked to original sources
PubMed1995

Association of p53 mutations with metastatic prostate cancer.

In prostate cancer, mutation of the p53 tumor suppressor gene has been associated with locally advanced disease and hormone-resistant disease that is predominantly localized to bone. However, little is known regarding the status of the p53 gene in metastatic prostate cancer that has not been treated with hormonal manipulation. We evaluated formalin-fixed, paraffin-embedded malignant tissues from 86 patients with various stages of prostate cancer, including pathologically confined, locally advanced, and metastatic disease, to detect abnormal p53 nuclear protein accumulation using immunohistochemistry. No abnormal p53 immunostaining was detected in 18 patients with prostate cancer confined to the gland. Two tumors from 21 patients with locally advanced disease (extracapsular extension and/or seminal vesicle invasion) had abnormal nuclear p53 accumulation, and a mutation in exon 7 of the p53 gene was detected in tumor DNA from one patient using single-strand conformation polymorphism-direct sequencing analysis. Of the remaining 47 patients studied in whom tissues from the prostate gland and a metastatic site (44 lymph node, 2 bone, and 1 lung) were available, only 3 had received hormonal therapy prior to obtaining metastatic tissue. In four patients both primary and metastatic tumors demonstrated accumulation of p53 protein, whereas seven additional patients exhibited p53 accumulation only at the metastatic site. In three patients the metastatic tumors harbored missense single-base substitutions in exon 5, as detected using single-strand conformation polymorphism-direct sequencing. These results indicate that p53 abnormalities are associated with lymph node metastases derived from prostate cancer patients that had not undergone hormonal therapy.

Adenocarcinoma↗
PubMed1994

Mesenchymal-epithelial interactions and transforming growth factor-beta expression during mouse prostate morphogenesis.

To explore the role of transforming growth factor-beta (TGF beta) isoforms and other growth-related genes during prostate morphogenesis in the mouse, we examined mRNA levels in fetal day 17 urogenital sinus, mesenchyme (UGM), and epithelium (UGE) as well as in the ventral, dorsal, and anterior lobes of the adult prostate. In addition, we used antiserum specific for extracellular TGF beta 1 in immunohistochemical studies to localize accumulation of the TGF beta 1 isozyme in the above tissues as well as those derived from fetal day 19 and neonatal mouse prostate. Differential patterns of expression in fetal and adult tissues were seen. TGF beta 1, -beta 2, and -beta 3 expression was substantially elevated in UGM compared to that in UGE, yet only TGF beta 1, not TGF beta 2 or TGF beta 3, mRNA levels were sustained in adult prostate tissues. High levels of accumulation of TGF beta 1 were demonstrated by immunohistochemistry in the mesenchymal compartment compared to those in the epithelial compartment throughout development. Interestingly, the highest levels of TGF beta 1 appeared in areas of active epithelial duct formation and delineated the mesenchymal architectural changes necessary for ductal network formation. Additional studies revealed that levels of mRNAs for other genes involved in tissue remodeling and growth were also elevated in UGM compared to those in UGE. Tissue plasminogen activator, urokinase plasminogen activator, androgen receptor, and c-myc mRNA levels were also elevated in UGM compared to UGE. Interestingly, whereas tissue plasminogen activator mRNA levels, like those of TGF beta 2 and -beta 3, were barely detectable in adult prostatic tissues, mRNA levels for urokinase plasminogen activator, androgen receptor, and c-myc were readily detected and expressed in a lobe-specific fashion. Overall, these data indicate that expression of TGF beta 1 isoforms and other growth-related genes is associated with mesenchymal cells in areas of active morphogenesis during prostate development and provide objective molecular and cellular information regarding mediators of mesenchymal-epithelial interactions in prostate.

Animals↗
PubMed1993

Transgenic models for the study of prostate cancer.

Transgenic model systems provide tools for obtaining information that clarifies important relationships between genetic alterations and carcinogenesis. One such relationship is the induction of specific growth factor activities by dominantly acting oncogenes. Using a "transgenic organ" model referred to as mouse prostate reconstitution (MPR) under conditions where the ras and myc oncogenes were introduced using a recombinant retrovirus into both the mesenchymal and epithelial compartments of the urogenital sinus, poorly differentiated prostate cancer (PC) was produced with high frequency (> 90%) in inbred C57BL/6 mice. Time-course studies using northern blotting and immunohistochemical analysis showed that the transition from benign to malignant status invariably was associated with the induction of elevated transforming growth factor-beta 1 (TGF-beta 1) expression. Additional immunohistochemical analysis of TGF-beta 1 in human PC and benign prostatic hyperplasia (BPH) showed that positive extracellular staining was significantly more extensive in PC compared with BPH. This differential staining pattern was evident in focal areas of PC adjacent to BPH. These findings in both the MPR model system and human PC suggest that elevated TGF-beta 1 expression is involved in the progression to malignancy and that its pattern of expression may become a useful marker of PC. Additional studies using transgenic animal models will continue to provide important clinically useful information about PC in man.

Animals↗
PubMed1993

Genetic predisposition and mesenchymal-epithelial interactions in ras+myc-induced carcinogenesis in reconstituted mouse prostate.

Using a mouse prostate reconstitution (MPR) model system, strain-specific responses to the ras and myc oncogenes were investigated. When ras + myc were introduced into both the mesenchymal and epithelial compartments of the urogenital sinus, poorly differentiated prostate cancer was produced at a high frequency (> 90%) in inbred C57BL/6 mice. In contrast, under similar conditions, inbred BALB/c MPRs formed benign prostatic hyperplasia that converted to cancer at a low frequency (< 10%). Restricting the oncogenes to the mesenchymal or epithelial compartments revealed that oncogene activities were more pronounced in the mesenchyme of C57BL/6 mice and resulted in elevated transforming growth factor-beta 1 expression along with a severe desmoplastic reaction. Heterologous MPRs composed of BALB/c mesenchyme and C57BL/6 epithelium or vice versa demonstrated that intrinsic properties of BALB/c mesenchyme can arrest the progression of ras + myc-initiated C57BL/6 epithelium from benign hyperplasia to malignant carcinoma.

Animals↗
PubMed1992

Transforming growth factor beta 1 as a biomarker for prostate cancer.

Using the mouse prostate reconstitution (MPR) model system, under conditions where the ras and myc oncogenes are introduced via a recombinant retrovirus into both the mesenchymal and epithelial compartments of the urogenital sinus, poorly differentiated prostate cancer is produced with high frequency (> 90%) using inbred C57BL/6 mice. Northern blotting and immunohistochemical analysis showed that the transition from benign prostatic hyperplasia (BPH) to prostate cancer is invariably associated with the induction of elevated transforming growth factor-beta 1 (TGF-beta 1) expression. Similar analysis of TGF-beta 1 in human BPH and prostate cancer is consistent with our MPR results and indicates that the accumulation of extracellular TGF-beta 1 is significantly more intense in prostate cancer compared to normal or benign prostate tissues. Interestingly, where benign pathologies are observed in the prostatic stroma in the presence of benign prostatic epithelium, extracellular TGF-beta 1 is seen predominantly in the stromal compartment. Experimental studies clearly demonstrate that mRNA levels of TGF-beta 1 and other growth related genes are regulated by androgens in prostate cancer cells. Overall, our results suggest that elevated TGF-beta 1 is involved in the development of prostate cancer. Direct determination of TGF-beta 1 levels and distribution as well as analysis of localized and systemic effects produced by TGF-beta 1 may serve as useful biomarkers for prostate cancer.

Animals↗
PubMed1992

Androgen sensitivity and gene expression in ras + myc-induced mouse prostate carcinomas.

We established an androgen-sensitive cell line (BR31-5) from a ras + myc-induced mouse prostate carcinoma and used this cell line together with a previously reported transplantable androgen-independent mouse prostate carcinoma to investigate patterns of expression for apoptosis-related genes in an androgen-deprived environment. Single cell suspensions derived from the BR31-5 cell line were inoculated into the flank of intact or castrated adult male C57BL/6 mice and tumors were harvested 12 days post-inoculation for Northern blotting. A transplantable androgen-independent prostate cancer was also inoculated into intact or castrated mice and tumors harvested 21 days later. Tumor volume analyses showed that BR31-5 carcinomas were androgen-sensitive. Northern blotting showed that mRNA levels for two apoptosis-related genes, transforming growth factor-beta 1 and c-myc, were significantly elevated to a similar extent in carcinomas grown in castrated hosts compared to intact hosts for both the androgen-sensitive BR31-5 and androgen-independent carcinomas. Levels of mRNA for tissue type plasminogen activator, shown previously to be elevated in androgen-independent carcinomas following growth in castrates, were also increased in BR31-5 carcinomas under similar androgen-deprived conditions but to a lesser extent. Interestingly, testosterone repressed prostate mRNA No. 2 levels shown previously to be similar in both the intact and castrated groups for androgen-independent carcinomas were significantly increased in the castrated group compared to the intact group for BR31-5 carcinomas. Therefore, specific patterns of expression for apoptosis-related genes may be able to discriminate androgen-sensitive and androgen-independent prostate cancer under androgen-deprived conditions.

Androgens↗
PubMed1991

Elevated transforming growth factor-beta 1 and beta 3 mRNA levels are associated with ras + myc-induced carcinomas in reconstituted mouse prostate: evidence for a paracrine role during progression.

Mouse prostate reconstitution is a useful model for studying the progression of ras + myc-induced carcinomas. When these oncogenes were introduced into both the epithelial and the mesenchymal compartments, poorly differentiated adenocarcinomas resulted. Restricted introduction of both oncogenes into the epithelium produced epithelial hyperplasia. Malignancies were produced in two out of 17 cases of selectively transformed epithelium, suggesting that the hyperplastic condition represents a premalignant phenotype. Restricted introduction of both oncogenes into the mesenchyme produced only mesenchymal dysplasia. Transforming growth factor-beta 1 (TGF-beta 1) and beta 3 (TGF-beta 3) mRNA levels were elevated in the ras + myc-induced carcinomas when compared to the normal controls or to the epithelial hyperplasias. In contrast, TGF-beta 2 mRNA levels were similar in all control and ras + myc-induced carcinomas. Elevated TGF-beta 1 mRNA levels were also found in mesenchymal dysplasia pointing to a potential paracrine activity by the ras + myc transformed mesenchyme. We conclude that elevated TGF-beta 1 and beta 3 are correlated with progression to malignancy and that mesenchyme derived TGF-beta 1 may play an important role in the promotion of ras + myc-induced carcinomas in this model system.

Animals↗
PubMed1991

Experimental oncogene induced prostate cancer.

The mouse prostate reconstitution model exploits the ability of the fetal urogenital sinus to differentiate into a mature prostate when grafted under the renal capsule of an adult isogenic male host. By use of a recombinant retroviral vector, the ras and myc oncogenes are introduced singly or in combination into the fetal urogenital sinus--resulting in distinct phenotypes of prostatic pathology: dysplasia (caused by ras), hyperplasia (caused by myc) and frank carcinomas (caused by a combination of ras+myc). This unique experimental model creates in vivo conditions that mimic the natural initiation and progression of cancer. An expanded MPR protocol allows restricted retrovirus infection of the mesenchyme or epithelial compartments to evaluate paracrine activities. It enables almost unparalleled flexibility in addressing fundamental questions in prostate cancer. We have identified genetic variance in the susceptibility to tumour induction between two different strains of mice (mimicking the observation of racial variability in the predisposition to clinical prostate cancer). The MPR model supports data from other tumour models and implicates TGF-beta 1 and TGF-beta 3 as being strongly associated with tumour progression. Finally, with this model, we have established clonal prostate adenocarcinomas to study directly the affects of castration on gene expression. Not only are TGF-beta 1 and TGF-beta 3 mRNA levels increased in association with malignancy but they are also further enhanced by castration treatment. Based on these experimental studies, we believe that TGF-beta 1 and TGF-beta 3 expression strongly influences the progression of prostate cancer. This information will hopefully impact on the development of more effective therapy for this important malignancy.

Adenocarcinoma↗
PubMed1990

Expression of the cDNA for the beta subunit of human casein kinase II confers partial UV resistance on xeroderma pigmentosum cells.

An immortalized xeroderma pigmentosum cell line belonging to the complementation group D (XP-D) was transfected with a normal human cDNA clone library constructed in a mammalian expression vector. Following UV-irradiation-selection, a transformant having a stable, partially UV-resistant phenotype was isolated. A transfected cDNA of partial length was rescued from the transformant's cellular DNA by in vitro amplification, using expression-vector specific oligonucleotides as primers in a polymerase chain reaction (PCR). Expression of this cDNA complemented the UV sensitivity of the XP-D cell line to the UV-resistance levels characteristic of the primary transformant. The nucleotide sequence of the cDNA was determined. The deduced protein identified the cDNA as encoding for the beta subunit of casein kinase II (CKII-beta). Similar to the effect exerted by the truncated CKII-beta cDNA, expression of a cDNA clone encompassing the complete translated region of CKII-beta leads to XP-D cells partially resistant to UV-irradiation. However, transfection of CKII-beta cDNA could also partially complement the UV-sensitivity of a xeroderma pigmentosum cell line belonging to group C (XP-C). Analysis by Southern, Northern and RNAase mismatch cleavage techniques did not reveal any functional defect in the CKII-beta gene of cell lines derived from either 7 XP-D or 10 XP-C families. We therefore consider it unlikely that either the XP-D or the XP-C DNA repair deficiency is associated with a defect in the beta subunit of casein kinase II. Nevertheless, our findings suggest the possibility that the cell's response to DNA damage is modulated by CKII-dependent protein phosphorylation.

Base Sequence↗
PubMed1989

Two new members of the OmpR superfamily detected by homology to a sensor-binding core domain.

The OmpR superfamily includes proteins that act as transcriptional regulators of operons that respond to environmental stimuli. A homologous domain near the N-terminus, termed a sensor-binding core domain, is thought to play a role in recognition of a signal transduction protein. We have identified two previously unrecognized members of this regulator family of proteins: a 23.8-kd protein transcribed from the uvrC transcription unit and the PgtA gene product, which is a phosphoglycerate transport regulatory protein. The sensor-binding core domain is also present in four proteins that regulate bacterial sporulation and chemotaxis. The 23.8-kd protein also has sequence similarity to elongation factor Tu and two regulatory proteins: HtpR, the heat-shock regulatory protein, and TraJ, a regulator of expression of genes involved in conjugation. There is a 77-amino acid region near the C-terminus of the 23.8-kd protein that has 30% similarity with a 28.1-kd protein coded for by an open reading frame 5' to the reading frame of the 23.8-kd protein in the uvrC transcription unit. Genetic distance analysis of amino acid sequences of proteins with a sensor-binding core domain suggests that the 23.8-kd protein and the chemotaxis regulatory proteins are distantly related to the other regulatory proteins in the OmpR superfamily.

Amino Acid Sequence↗
PubMed1989

Intermolecular plasmid recombination in fibroblasts from humans with DNA damage-processing defects.

We have evaluated the ability of immortalized human fibroblasts to recombine transfected plasmid DNA. A number of cell lines from normal individuals and from patients with DNA damage-processing defects were examined. Two plasmid recombination substrates were derived from pSV2neo and contained nonoverlapping deletions in the aminoglycoside phosphotransferase II gene. Intermolecular recombination was assessed by two methods after cotransfection. In a short-term, extrachromosomal recombination assay, low molecular weight DNA was extracted from the human cells 48 h after transfection, and recombinant plasmids were detected by transformation into appropriate indicator bacteria. In a long-term stable recombination assay the fibroblasts were cotransfected and G418-resistant colonies allowed to form. By the former assay all but two cultures were recombination-proficient, whereas all were recombination-proficient by the latter assay. The efficiency of transfection of human cells with plasmids appears to be a major variable affecting recombination. Recombination can be stimulated by uv irradiation of plasmid DNA prior to transfection. Cells from patients with Fanconi anemia, ataxia telangiectasia, and xeroderma pigmentosum complementation groups A, C, D, E, and G are not defective at intermolecular plasmid recombination.

Cell Line↗
PubMed1988

Diseases with DNA damage-processing defects.

Xeroderma pigmentosum, Cockayne's syndrome, ataxia telangiectasia, Fanconi anemia, and Bloom's syndrome are autosomal recessive diseases with cellular defects in the ability to process DNA damage. Although these diseases are rare, they are seen occasionally in practice and provide insight into the mechanisms of DNA repair and replication in humans. The authors will review the clinical and cytological presentation of each disease, the genetic heterogeneity, as inferred by complementation analysis, and the differentiating characteristics of each. The authors will conclude with a discussion of the state of current research on each disease and possible directions for future research.

Anemia, Aplastic↗
PubMed1987

Transformation of DNA repair-deficient human diploid fibroblasts with a simian virus 40 plasmid.

Fibroblasts from patients with xeroderma pigmentosum (XP) complementation groups A, C, D, E, and G, as well as Bloom syndrome (BS) and Fanconi anemia (FA) have been transfected with a plasmid, pSV7, containing the early region of Simian virus 40 (SV40). All of the cultures exhibited cytologic changes characteristic of transformed cells and expressed T-antigen. They also contained integrated copies of DNA derived from the vector, and in several cases, extrachromosomally replicated DNA. Not all of the transfected cultures became immortalized. The transformed xeroderma pigmentosum (XP) cultures retained their UV-sensitive phenotype in all but one case. The BS and FA cell lines retained their characteristic phenotype. All of the cultures, except the BS cells, can be readily transfected with the plasmids, pSV2neo and pSV2gpt.

Bloom Syndrome↗
PubMed1987

DNA polymerase III of Escherichia coli is required for UV and ethyl methanesulfonate mutagenesis.

Strains of Escherichia coli possessing the pcbA1 mutation, a functional DNA polymerase I, and a temperature-sensitive mutation in DNA polymerase III can survive at the restrictive temperature (43 degrees C) for DNA polymerase III. The mutation rate of the bacterial genome of such strains after exposure to either UV light or ethyl methanesulfonate was measured by its rifampicin resistance or amino acid requirements. In addition, Weigle mutagenesis of preirradiated lambda phage was also measured. In all cases, no increase in mutagenesis was noted at the restrictive temperature for DNA polymerase III. Introduction of a cloned DNA polymerase III gene returned the mutation rate of the bacterial genome as well as the Weigle mutagenesis to normal at 43 degrees C. Using a recA-lacZ fusion, the SOS response after UV irradiation was measured and found to be normal at the restrictive and permissive temperature for DNA polymerase III, as was induction of lambda prophage. Recombination was also normal at either temperature. Our studies demonstrate that a functional DNA polymerase III is strictly required for mutagenesis at a step other than SOS induction.

DNA Polymerase III↗
PubMed1984

Induction of bacteriophage from members of the Mycobacterium avium, Mycobacterium intracellulare, Mycobacterium scrofulaceum serocomplex.

Bacteriophages have been induced from strains in the Mycobacterium avium, Mycobacterium intracellulare, Mycobacterium scrofulaceum serocomplex by exposure of cultures to UV light or treatment with mitomycin C. One-sixth of the strains examined, representing all but one of the 31 authenticated serotypes, were found to possess phages lytic for a Mycobacterium smegmatis indicator strain. Four single-plaque isolated phages, TM4, TM9, TM10 and TM20, were purified and shown to have a similar morphology on electron micrographs. They had an isometric head of diameter 50-58 nm and a flexible non-contractile tail about 170 nm in length with a terminal bulb. All had an identical buoyant density in CsCl of 1.521 g cm-3 and extreme sensitivity to chloroform. The induced phages differed in host range and possessed the unique ability to lyse other members of the serocomplex. Interest in these phages centres on a possible role in mediating genetic interrelationships between members of the serocomplex.

Lysogeny↗
PubMed1981

General Transduction in Vibrio cholerae.

Evidence was obtained for general transduction in Vibrio cholerae. Transduction of three amino acid markers and three antibiotic resistance characters was demonstrated using strains of biotype eltor and biotype cholerae. Some of the genetic characters were transduced from a biotype eltor donor (and its mutant derivatives) to biotype cholerae and eltor recipients. For the genetic traits examined, the frequencies of transduction ranged between 10(-5) and 10(-8). Maximal frequencies were obtained with transducing phage lysates that were irradiated with ultraviolet light. The development of a system of general transduction will now aid in fine structure analysis and detailed mapping of the chromosome of V. cholerae.

Bacteriophages↗
PubMedDate not supplied

Cooperative therapeutic effects of androgen ablation and adenovirus-mediated herpes simplex virus thymidine kinase gene and ganciclovir therapy in experimental prostate cancer.

Adenovirus-mediated transduction of the herpes simplex thymidine kinase gene (HSV-tk) in conjunction with ganciclovir (GCV) has been shown to result in significant growth suppression and to enhance survival in a model of mouse prostate cancer. However, this therapeutic activity is not sustained, because in most cases tumors eventually regrow and ultimately cause the death of the host. Androgen ablation, an inducer of apoptosis in prostate cells which is used widely as palliative therapy in patients with prostate cancer, was combined with HSV-tk plus GCV using an androgen-sensitive mouse prostate cancer cell line. The combination of castration and HSV-tk plus GCV led to markedly enhanced tumor growth suppression in both subcutaneous and orthotopic models compared with either treatment alone and resulted in an enhanced survival in which combination-treated animals lived twice as long as controls in the subcutaneous model and over 50% longer than controls in the orthotopic model. Further analysis of apoptotic activity demonstrated high levels of apoptosis only in combined androgen ablation and HSV-tk plus GCV-treated tumors after 14 days of growth in an androgen-depleted environment and 8 days after HSV-tk plus GCV therapy. At this time, the apoptotic index, but not the percent of necrotic tissue, was significantly higher for combination therapy-treated tumors relative to control-treated tumors or either treatment alone. These data indicate that the therapeutic effects of androgen ablation and HSV-tk plus GCV are cooperative and that increased apoptosis may, in part, underlie these activities.

Adenoviridae↗