Biomedical subjects

J C Romijn

Publications and source records attributed to J C Romijn.

At least 37 records · Page 2Linked to original sources

PubMed1998

Increased calcium oxalate monohydrate crystal binding to injured renal tubular epithelial cells in culture.

The retention of crystals in the kidney is considered to be a crucial step in the development of a renal stone. This study demonstrates the time-dependent alterations in the extent of calcium oxalate (CaOx) monohydrate (COM) crystal binding to Madin-Darby canine kidney (MDCK) cells during their growth to confluence and during the healing of wounds made in confluent monolayers. As determined by radiolabeled COM crystal binding studies and confirmed by confocal-scanning laser microscopy, relatively large amounts of crystals (10.4 +/- 0.4 micrograms/cm2) bound to subconfluent cultures that still exhibited a low transepithelial electrical resistance (TER < 400 omega.cm2). The development of junctional integrity, indicated by a high resistance (TER > 1,500 omega.cm2), was followed by a decrease of the crystal binding capacity to almost undetectable low levels (0.13 +/- 0.03 microgram/cm2). Epithelial injury resulted in increased crystal adherence. The highest level of crystal binding was observed 2 days postinjury when the wounds were already morphologically closed but TER was still low. Confocal images showed that during the repair process, crystals selectively adhered to migrating cells at the wound border and to stacked cells at sites were the wounds were closed. After the barrier integrity was restored, crystal binding decreased again to the same low levels as in undamaged controls. These results indicate that, whereas functional MDCK monolayers are largely protected against COM crystal adherence, epithelial injury and the subsequent process of wound healing lead to increased crystal binding.

Animals↗

PubMed1997

Orthotopic implantation of human prostate cancer cell lines: a clinically relevant animal model for metastatic prostate cancer.

BACKGROUND: To study the metastatic behavior of human prostate cancer cell lines, orthotopic injection in nude mice was performed. METHODS: Local tumor growth, metastasis formation, prostate-specific antigen, and androgen receptor expression were examined. RESULTS: Hormone-independent cell lines (PC-3, PC-3-125-IL, and TSU-Pr1) were highly tumorigenic and had a higher rate of lymph node metastasis after orthotopic than after subcutaneous implantation. PC-3 cell lines also consistently metastasized to the lungs. The androgen-sensitive LNCaP cell line showed local growth in 7 of 10, and lymph node metastasis in 4 animals. Significant serum PSA levels and strong receptor expression in primary and metastatic tumor tissues were observed. CONCLUSIONS: These results demonstrates that orthotopic implantation of human prostate cancer cell lines, including LNCaP, reproducibly leads to metastasis formation in nude mice.

Animals↗

PubMed1997

Stromal inhibition of epithelial cell growth in the prostate; overview of an experimental study.

The paracrine influence of prostatic stroma on the proliferation of prostatic epithelial cells was investigated. Using a double-layer soft agar assay it was demonstrated that stromal cells from the human prostate inhibit the anchorage-independent growth of the prostatic tumor epithelial cell lines PC-3 and LNCaP. Anchorage-dependent growth was inhibited too as was shown in the semi-automated colorimetric MTT test performed on multiwell plates. Antiproliferative activity was mediated by a diffusible factor in the stromal cell conditioned medium and was found to be produced specifically by prostatic stromal cells. Although the putative inhibiting factor shared some properties with transforming growth factor beta (TGF-beta) evidence is presented that the factor is different from this well-known inhibitor of epithelial cell growth. Absence of TGF-beta activity was shown by the lack of inhibitory response of the TGF-beta-sensitive mink lung cell line CCL-64 to prostate stromal cell conditioned medium and to concentrated partially purified preparations of the inhibitor. Furthermore, neutralizing antibodies against TGF-beta 1 or TGF-beta 2 did not cause a decline in the level of PC-3 growth inhibition caused by partially purified inhibitor. It is concluded that the prostate stroma-derived factor may be a novel growth inhibitor different from any of the currently described inhibiting factors.

Antibodies, Blocking↗

PubMed1997

Clinical usefulness of RT-PCR detection of hematogenous prostate cancer spread.

Understaging is commonly associated with therapeutic failure of surgical intervention in apparently localized prostate cancers. Methods that specifically detect prostate cancer cells in the circulation may be able to identify metastatic cancers and thus aid in the selection of the most adequate therapy. The high sensitivity and specificity of the reverse transcriptase-polymerase chain reaction (RT-PCR) encouraged various groups to investigate the mRNA expression of prostate-specific markers in the peripheral blood of patients with prostate cancer. However, probably due to methodological differences, many contradictory results have been obtained with the markers studied so far: prostate-specific antigen (PSA) and prostate-specific membrane antigen (PSM). For this reason, clinical decisions should not be based yet on RT-PCR results. Future research and long-term follow-up on the patients may point out whether RT-PCR assays, following appropriate standardization, will have an additive value in prostate cancer staging and in prediction of tumor progression.

Antigens, Surface↗

PubMed1997

Cell cultures and nephrolithiasis.

While the physical chemistry of stone formation has been intensively studied during the last decade, it has become clear that the pathophysiology of renal stone disease cannot be explained by crystallization processes only. In recent years, evidence has emerged that the cells lining the renal tubules can have an active role in creating the conditions under which stones may develop. Since it is difficult to study these mechanisms in vivo, cultured renal tubular cells have become increasingly popular for the study of physiological and cell biological processes that are possibly linked to stone disease. In this paper, we discuss the possible contribution of cellular processes such as transepithelial oxalate transport and crystal--cell interaction to the formation of renal stones. Experimental studies that have been performed with cultured renal cells to elucidate the mechanisms involved in these processes will be summarized.

Animals↗

PubMed1997

Decreased levels of topoisomerase II alpha in human renal cell carcinoma lines resistant to etoposide.

Renal cell carcinoma (RCC) displays strong resistance against many chemotherapeutic drugs. Overexpression of P-glycoprotein (Pgp) appears to be part of this resistance. The involvement of another resistance mechanism, involving the decreased activity of DNA topoisomerase II (topoII), remains uncertain. By culturing the human RCC lines RC2 and RC21 in the presence of increasing concentrations of etoposide, we derived the variant sublines RC2E, RC21A and RC21E, that had acquired approximately 30-, 60- and 90-fold resistance to this drug respectively. RC2E, RC21A and RC21E were approximately 50-, 5- and 400-fold cross-resistant to doxorubicin respectively. RC2E and RC21E also showed cross-resistance (approximately 200- and 3500-fold respectively) to vinblastine. Quantitative differences in MDR1 and Pgp expression (elevated in RC2E and RC21E) and topoII alpha (reduced in RC21E and RC21A) were demonstrated using Western blotting and the reverse transcriptase/polymerase chain reaction. Decreased amounts of topoII alpha were reflected in a reduced activity of RC21A and RC21E as measured by unknotting phage P4 DNA. Qualitative changes of the topoII alpha gene, such as point mutations in the motif B/DNBS and DNA-binding regions, or differences in methylation status of the promoter gene of RC21E, were not found. These cell lines represent a model of a solid tumor in which overexpression of Pgp, a combination of increased Pgp and decreased topoII alpha, and a decrease of topoII alpha are represented.

Antigens, Neoplasm↗

PubMed1997

Specific cytogenetic aberrations in two novel human prostatic cell lines immortalized by human papillomavirus type 18 DNA.

Using chromosome banding and fluorescence in situ hybridization (FISH) with painting probes, sequential cytogenetic analysis was performed of two novel prostate cell lines, PZ-HPV-7 and CA-HPV-10, established by human papillomavirus (HPV) 18 DNA transformation. PZ-HPV-7 originates from a normal diploid prostate epithelial cell strain. PZ-HPV-7 progressed from an initial diploid to a hypertetraploid chromosome number with a relative gain of chromosomes 5 and 20 (7 to 8 copies each). Structural changes were limited; 3p- (2 copies), 3q- (1 copy), and possibly a der(16p;12q). CA-HPV-10 originates from an epithelial cell strain derived from a high-grade human prostate cancer specimen, which showed several karyotypic abnormalities including an extra Y chromosome and double minutes (dmin). In early passage the karyotype of CA-HPV-10 appeared unstable with a decreasing number of cells exhibiting dmin. In late passage the dmin were replaced by a large homogeneously staining region (hsr) on 9p+ marker. The hsr was shown by FISH to be of chromosome 1 origin. The modal number was mainly hypertriploid (72, range 69 to 75). Loss of Y was remarkable (0 to 1 copy). Consistent markers included two copies each of del(1)(q12q31) and der(9)t(1;9)(?;p22), and one der(11)t(4;11) (?;q21). HPV type 18 genomic integration sites were identified on 1p for PZ-HPV-7 and on the 9p+ marker for CA-HPV-10. In conclusion, both PZ-HPV-7 and CA-HPV-10 showed clonal cytogenetic changes. These two cell lines constitute a novel in vitro model to study the mechanisms involved in human prostate carcino-genesis.

Cell Line, Transformed↗

PubMed1997

Does urinary oxalate interfere with the inhibitory role of glycosaminoglycans and semisynthetic sulfated polysaccharides in calcium oxalate crystallization?

OBJECTIVES: Previously it was shown that the polysaccharide G872 in vitro strongly inhibits calcium oxalate monohydrate crystallization processes. However, when rats on a stone-inducing diet of ethylene glycol plus vitamin D3 are given this polysaccharide, no changes in the urine capacity for crystallization inhibition were found. We investigated here how the inhibitory action of polysaccharides changes under high oxalate conditions, as they exist in the stone inducing diet. METHODS: Calcium oxalate monohydrate (COM) crystals were incubated in a series of 0.05 M PBS buffers containing polysaccharides with increasing oxalate concentrations (0-0.4 mmol/l). The coated crystals were collected, washed and resuspended in an artificial urine. We then measured the zeta potential of the crystals, using a Coulter DELSA 440, and the initial rates for crystal growth and agglomeration, using the Coulter Multisizer II. RESULTS: Addition of oxalate to the medium shifts the negative zeta potential distribution of COM crystals coated by polysaccharides in positive direction. Particle size analysis demonstrated that the initial rates of COM crystal growth and agglomeration responding to oxalate concentration changes (0.1-->0.4 mmol/l) in the presence of G872 (0.2 mg/l) are approximately 2.5 times faster than that in the absence of G872. CONCLUSIONS: Oxalate interferes with the binding of polysaccharides to crystals. This can be envisioned to occur through changes in the crystal surface properties or by induction of functional and secondary structural changes of urinary macromolecular inhibitors such as GAGs, resulting in a decrease of their inhibitory activity against COM crystallization. Thus, in urine, a high oxalate may increase the rate of crystallization both by increasing the supersaturation and by decreasing the inhibitory potential of the urine.

Calcium Oxalate↗

PubMed1996

Tissue specific and androgen-regulated expression of human prostate-specific transglutaminase.

Transglutaminases (TGases) are calcium-dependent enzymes catalysing the post-translational cross-linking of proteins. In the prostate at least two TGases are present, the ubiquitously expressed tissue-type TGase (TGC), and a prostate-restricted TGase (TGP). This paper deals with the molecular cloning and characterization of the cDNA encoding the human prostate TGase (hTGP). For this purpose we have screened a human prostate cDNA library with a probe from the active-site region of TGC. The largest isolated cDNA contained an open reading frame encoding a protein of 684 amino acids with a predicted molecular mass of 77 kDa as confirmed by in vitro transcription-translation and subsequent SDS/PAGE. The hTGP gene was tissue-specifically expressed in the prostate, yielding an mRNA of approx. 3.5 kb. Furthermore, a 3-fold androgen-induced upregulation of hTGP mRNA expression has been demonstrated in the recently developed human prostate cancer cell line, PC346C. Other well established human prostate cancer cell lines, LNCaP and PC-3, showed no detectable hTGP mRNA expression on a Northern bolt. The gene coding for prostate TGase was assigned to chromosome 3.

Amino Acid Sequence↗

PubMed1996

Oxalate transport and calcium oxalate renal stone disease.

Hyperoxaluria is considered to play a crucial role in calcium oxalate (CaOx) renal stone disease. The amount of oxalate excreted into the urine depends on intestinal absorption, endogenous production, renal clearance and renal tubular transport. Since a primary disorder has not been found so far in most CaOx stone formers and since oxalate is freely filtered at the glomerulus, most studies are presently focussed on alterations in epithelial oxalate transport pathways. Oxalate can be transported across an epithelium by the paracellular (passive) and transcellular (active) pathway. Oxalate transport across cellular membranes is mediated by anion-exchange transport proteins. A defect in the structure of these transport proteins could explain augmented transcellular oxalate transport. Little is known about the physiological regulation of oxalate transport. In this review cellular transport systems for oxalate will be summarized with special attention for the progress that has been made to study oxalate transport in a model of cultured renal tubule cells. Better understanding of the physiological processes that are involved in oxalate transport could yield information on the basis of which it might be possible to design new approaches for an effective treatment of CaOx stone disease.

Animals↗

PubMed1996

Gain and loss of chromosomes 1, 7, 8, 10, 18, and Y in 46 prostate cancers.

Fluorescence in situ hybridization (FISH) with centromere probes was used to investigate numerical aberrations of chromosomes 1, 7, 8, 10, 18, and Y in 46 prostate carcinoma (PC) and 11 benign prostatic hyperplasia (BPH) samples. None of the benign specimens showed any chromosomal aberration. Forty-one of 46 PC specimens showed numerical aberrations of one or more chromosomes. All investigated chromosomes showed numerical aberrations in at least 30% of the specimens, gain being more frequent than loss. Comparison of DNA flow cytometry (FCM) and FISH results showed that not only aneuploid tumors but also most diploid tumors harbored numerical chromosome aberrations. Chromosome 10 was the most frequently gained (65%), and Y the most frequently lost chromosome (14%). Nonmetastatic and metastatic tumors differed significantly (P < .05) in the number of copies for chromosomes 7, 8, and 10, but not for 1, 18, and Y. These results suggest strongly that gains of chromosomes 7, 8, and 10 are involved in PC progression.

Aged↗

PubMed1996

Crystal-cell interaction inhibition by polysaccharides.

PURPOSE: We studied the effect of polysaccharides on interactions between calcium oxalate monohydrate (COM) crystals and cultured renal cells. MATERIALS AND METHODS: Monolayers of Madin-Darby canine kidney (MDCK) cells were incubated with radiolabeled crystals in the presence of various concentrations of natural glycosaminoglycans (GAGs) and semisynthetic polysaccharides (SSPs). RESULTS: While most GAGs were found to have relatively little effect, SSPs (SP54, G871 and G872) were potent inhibitors of crystal-cell association. Pretreatment of crystals, but not of cells, was similarly effective, suggesting polysaccharide-induced modification of crystal surface properties. CONCLUSIONS: This result further supports the idea that SSPs, and especially G872, are of potential interest for treatment of recurrent stone disease.

Analysis of Variance↗

PubMed1996

Development of seven new human prostate tumor xenograft models and their histopathological characterization.

Seven human prostate tumor models were established by transplanting tumor fragments in NMRI athymic nude mice. Once established, the tumors were serially transplantable in both NMRI and BALB/c nude mice. The xenografts originated from primary prostatic carcinomas (prostatectomy specimens), transurethral resection material, and metastatic lesions (pelvic lymph nodes and scrotal skin). Histological examination revealed that, in the course of several mouse passages (8 to 23), tumors retained their resemblance to the original patient material. The PC-295, PC-310, PC-329, and PC-346 tumors are dependent on androgens for their growth. The PC-324, PC-339, and PC-374 tumors are androgen independent, although growth of PC-374 tumors still seemed androgen sensitive. All tumors are diploid, except for the PC-374, which is tetraploid. The diploid PC-295 tumor has an additional small population of tetraploid cells. All xenografts displayed a heterogeneous expression pattern of the androgen receptor except for the PC-324 and PC-339 tumors in which the androgen receptor could not be detected. Prostatic acid phosphatase and prostate-specific antigen were retained during serial transplantation in all tumors but the PC-324 and PC-339. This panel of permanent human prostate tumor models comprises tumors representing both the androgen-dependent and -independent stages of human prostate cancer with various degrees of differentiation and, therefore, is of great value for the study of many aspects of growth and progression of human prostate cancer.

Animals↗

PubMed1996

Zeta potential measurement and particle size analysis for a better understanding of urinary inhibitors of calcium oxalate crystallization.

To better understand urinary inhibitors of calcium oxalate crystallization, both zeta potential measurement and particle size analysis were chosen to illustrate: (1) the potential therapeutic efficacy of G872, a semi-synthetic sulfated polysaccharide, in stone prevention; and (2) the relative contribution of various urinary fractions ¿e.g., ultrafiltered urine (UFU), Tamm-Horsfall protein (THP), urinary polyanions precipitated with cetylpyridinium chloride (CPC), urinary macromolecular substances with different concentration ratios (UMS10,50,90 and UMS'10,50,90) and THP-free urine (THPFU)¿ to total urinary inhibitory activity. The results showed: (1) addition of G872 significantly enhances urinary inhibitory activity and negative zeta potential values; (2) re-addition of the CPC to UFU completely restores urinary inhibitory activity; and (3) artificial urines prepared by mixing UMS'10,50,90 from THPFU with UFU differed in inhibitory activity from that prepared by mixing UMS10,50,90 from a pooled normal urine with UFU. Based on these experimental results, the following speculations can be made: (1) normal human urines are considered to be a protective colloidal system; (2) urinary inhibitory activity originates mainly from CPC and/or UMS; (3) normal THP is a protective material to maintain urinary inhibitory activity; and (4) mutual interaction between urinary inhibitors may change the total urinary inhibitory activity.

Calcium Oxalate↗

PubMed1996

Lectin-cytochemistry of experimental rat nephrolithiasis.

Lectin reactivity in epithelial apical cell coats of normal rat kidneys was compared to that from animals subjected to crystal inducing diets (CID). The aim was to see whether the absence of lectin reactivity in cell coats is related to intratubular calcium oxalate crystal retention. In normal rat kidneys, after a pre-embedding procedure, it was observed that at the ultrastructural level, reactivity was present but that the lectin specificity for the various parts of the nephron might have to be reconsidered. There was heterogeneity between the epithelial cells with respect to the presence of coat material in the tubular cell apices. Tubular epithelial cell apices from CID rats showed no obvious changes in lectin reactivity pattern. Lectin reactivity was present at the periphery of intratubular crystals but undetectable at true crystal attachment sites or reduced at cell apices in the vicinity of recently attached crystals or agglomerates. After a post-embedding reaction procedure, wheat-germ agglutinin (WGA)-lectin reactivity confirmed the presence of coat material in the cleft between cell apex and retained crystal at crystal-attachment sites. The WGA/Au-10 nm reaction products were also seen inside epithelial cells. WGA/Au-10 nm reaction products mark a crystal matrix component inside intratubular and retained crystals. A similar matrix was also marked by an alpha-osteopontin (alpha OPN/Au-10 nm) reaction product.

Animals↗

PubMed1995

Inhibition of prostatic epithelial cell proliferation by a factor secreted specifically by prostatic stromal cells.

Stromal cells from the prostate were recently shown to inhibit clonal growth of the prostatic carcinoma cell lines PC-3 (hormone-independent) and LNCaP (hormone-sensitive) in coculture. Our study revealed that stromal cell-conditioned medium strongly inhibited proliferation of PC-3 and LNCaP cells when grown in monolayer culture. Antiproliferative activity was found to be reversible, and was produced specifically by prostatic stromal cells and not by stromal cells derived from skin, foreskin, uterus, kidney, and Wilms' tumor. Inhibition was not species-specific, since the cell lines AT-2.1 and MATLyLu, derived from the Dunning rat prostate tumor, were also sensitive. No inhibition, however, occurred on breast and renal carcinoma cell lines, suggesting a prostate-specific action. The putative inhibiting factor(s) could be concentrated and partially purified by ammonium sulfate precipitation. The possible role in stromal control of epithelial cell proliferation is discussed.

Animals↗

PubMed1995

Immunocytochemical characterization of explant cultures of human prostatic stromal cells.

The study of stromal-epithelial interactions greatly depends on the ability to culture both cell types separately, in order to permit analysis of their interactions under defined conditions in reconstitution experiments. Here we report the establishment of explant cultures of human prostatic stromal cells and their immunocytochemical characterization. As determined by antibodies to keratin and prostate specific acid phosphatase, only small numbers (< 5%) of epithelial cells were present in primary cultures; subsequent passaging further reduced epithelial cell contamination. Antibodies against intermediate filament proteins (keratins, vimentin, and desmin) and smooth muscle actin microfilaments demonstrated that stromal cells from benign prostatic hyperplasia and prostate carcinoma differed in regard to their differentiation markers. Two contrasting phenotypes were identified in cultures derived from these two different lesions: One exhibiting fibroblastic features, was predominant in cultures derived from benign lesions and a second, showing varying degrees of smooth muscle differentiation, was more abundant in carcinoma-derived cultures. These findings are indicative of a remarkable divergence in the stromal-epithelial relationships associated with these pathological conditions and may provide us with a potential tool for studying these processes.

Acid Phosphatase↗

PubMed1995

Stromal inhibition of prostatic epithelial cell proliferation not mediated by transforming growth factor beta.

The paracrine influence of prostatic stroma on the proliferation of prostatic epithelial cells was investigated. Stromal cells from the human prostate have previously been shown to inhibit anchorage-dependent as well as anchorage-independent growth of the prostatic tumour epithelial cell lines PC-3 and LNCaP. Antiproliferative activity, mediated by a diffusible factor in the stromal cell conditioned medium, was found to be produced specifically by prostatic stromal cells. In the present study the characteristics of this factor were examined. It is demonstrated that prostate stroma-derived inhibiting factor is an acid- and heat-labile, dithiothreitol-sensitive protein. Although some similarities with type beta transforming growth factor (TGF-beta)-like inhibitors are apparent, evidence is presented that the factor is not identical to TGF-beta or to the TGF-beta-like factors activin and inhibin. Absence of TGF-beta activity was shown by the lack of inhibitory response of the TGF-beta-sensitive mink lung cell line CCL-64 to prostate stromal cell conditioned medium and to concentrated, partially purified preparations of the inhibitor. Furthermore, neutralising antibodies against TGF-beta 1 or TGF-beta 2 did not cause a decline in the level of PC-3 growth inhibition caused by partially purified inhibitor. Using Northern blot analyses, we excluded the involvement of inhibin or activin. It is concluded that the prostate stroma-derived factor may be a novel growth inhibitor different from any of the currently described inhibiting factors.

Antibodies↗