Effect of oral cholecalciferol 2,000 versus 5,000 IU on serum vitamin D, PTH, bone and muscle strength in patients with vitamin D deficiency.

UNLABELLED: Treatment of vitamin D deficiency for 3 months with oral cholecalciferol 5,000 IU daily was more effective than 2,000 IU daily in achieving optimal serum 25-hydroxyvitamin D (25OHD) concentrations. Optimal 25OHD serum level calculated to be 63.8 nmol/L. All parameters of muscle strength improved following administration of cholecalciferol for 3 months. INTRODUCTION: The aim of this study was to determine the optimal dose of cholecalciferol required to achieve target serum 25OHD level ≥ 75 nmol/L and its relationship to both bone turnover and muscle strength. METHODS: Thirty deficient patients (serum 25OHD ≤ 50 nmol/L) were randomly assigned into two groups-i.e. 2,000 and 5,000 IU/day. Data were collected at baseline, at 2 and 3 months post-therapy: (a) clinical demographics, (b) dietary calcium recall, (c) physical tests of muscle function and (d) biochemistry. Statistical analysis used paired student t test and analysis of variance. Regression analysis was used to determine relationship between serum 25OHD and parathyroid hormone (PTH). RESULTS: Twenty-six (87%) patients completed 3 months of therapy. The percent increase in serum 25OHD (compared to baseline) was 82.7% in 2,000-IU group and 219.5% in 5,000-IU group. All participants (100%) achieved a serum 25OHD concentration >50 nmol/L; only 5 subjects (45.4%) in 2,000-IU group compared to 14 subjects (93.3%) in 5,000-IU group achieved final 25OHD concentration ≥ 75 nmol/L (p < 0.01). In the regression analysis, the reflexion point at which the PTH level increased above the normal range was calculated to be 63.8 nmol/L 25OHD. All parameters of muscle strength showed trends in improvements following the administration of both the 2,000 and 5,000 IU doses. No patient reported untoward side effects and no patient developed hypercalcaemia. CONCLUSION: Treatment for 3 months with oral cholecalciferol 5,000 IU daily may be more effective than 2,000 IU daily in achieving optimal serum 25OHD concentrations in vitamin D-deficient patients.

Extracellular matrix interactions. 1: Production of extracellular matrix with attachment and growth-sustaining functions by UWOV2 ovarian cancer cells growing in protein-free conditions.

Constitutive production of extracellular matrix with attachment and growth-promoting effects by an ovarian cancer cell line (UWOV2 (Pf)) growing in entirely protein-free conditions is described. This extracellular matrix has an ordered fibrillar, network structure consisting mainly of type IV collagen and laminin, as well as containing hyaluronan, glycoproteins, and proteoglycans. Type IV collagen appears to provide mainly structural support while other matrix components are responsible for the attachment and growth-promoting effects. This culture system provides an ideal model for studying the effects of extracellular matrix on cell attachment and growth. This system is also important in studying the concept of autonomous growth because the production of extracellular matrix by these cells appears to be growth regulatory even in an entirely protein-free culture system.

Extracellular matrix interactions. 2: Extracellular matrix structure is important for growth factor localization and function.

The influence of extracellular matrix components and of extracellular matrix structure on in vitro cell growth was investigated in the UWOV2 (Pf), protein-free cell culture model. This cell line constitutively produces an ordered extracellular matrix in the absence of any exogenous protein or growth factor. Extracellular matrix from UWOV2 (Pf) cells was found to contain both transforming growth factor beta (TGF beta) and platelet-derived growth factor (PDGF), which were shown to have an autostimulatory role for UWOV2 (Pf) cell growth. Matrix structure was shown to be important for allowing expression of the functional activity of these two growth factors. In addition, a nonuniform distribution of PDGF, embedded within the matrix structure, was demonstrated by immunoelectronmicroscopy. Apart from these two well-defined growth factors, additional but as yet unidentified growth stimulatory factor(s) were extractable from UWOV2 (Pf) extracellular matrix. These investigations indicate the potential role of extracellular matrix both as a mechanism for concentrating as well as modulating the function of cellular growth factors.

In vitro maintenance of a new ovarian cancer cell line in protein-free media: a potential model for autonomous growth and tumor progression.

A new cell line UWOV2 (pf) capable of long-term growth in the absence of any added serum protein, exogenous growth factor, insulin or transferrin, is described. The original cell line (UWOV2 and UWOV2 (sf), adapted to grow in serum-free conditions) was derived from the ascitic tumor of a patient with ovarian carcinoma. Under continuous culture conditions further adaptations have occurred enabling UWOV2 (pf) to maintain anchorage-dependent growth without requiring exogenous anchorage or growth factors. These cells produce a structured extracellular matrix which acts as an adhesive substrate for the UWOV2 (pf) cells themselves as well as for a number of other long-term cell lines including NRK and 3T3 cells. Furthermore, while UWOV2 (pf) cells produce a transforming growth factor beta (TGF beta)-like growth factor, they appear to be only partially dependent on autocrine growth stimulation, and other mechanisms for autonomous growth stimulation appear to exist. This cell line may be a useful model for the study of progressive growth autonomy in human tumors.

Interferon (IFN) alpha inhibits cell proliferation of UWOV2 cells without down-regulation of transferrin receptors.

It has been suggested that growth inhibition of cells by interferons may be mediated through interferon induced down-regulation of transferrin receptor expression. We describe a continuously growing cell line UWOV2 (pf) which expresses cell surface transferrin receptor but is able to grow in the absence of transferrin. This cell line is sensitive to the growth inhibitory effects of interferon alpha. Interferon alpha induced growth inhibition is not, however, accompanied by modulation of transferrin receptor expression suggesting that transferrin receptor modulation is not an essential component of the growth inhibitory effect of interferons.

Treatment of malignant ascites due to recurrent/refractory ovarian cancer: the use of interferon-alpha or interferon-alpha plus chemotherapy in vivo and in vitro.

Intraperitoneal treatment with interferon (IFN) for malignant ascites due to advanced ovarian carcinoma refractory to chemotherapy gave an objective response rate of 36% (7/19 patients treated). In vitro studies demonstrated that cytotoxicity of peripheral blood monocytes/macrophages was stimulated by IFN. However, peritoneal exudate cells obtained after intraperitoneal treatment with interferon were not stimulated to kill autologous tumour cells. Clinical response was therefore most probably due to a direct inhibitory effect of IFN on growth of malignant cells rather than due to an immune modulatory effect. Using a newly established ovarian cancer cell line (UWOV1), synergy between the growth inhibitory/antitumour effects of IFN and cisplatin was demonstrated at clinically achievable concentrations of each agent. IFN plus cisplatin proved to be more effective than intraperitoneal cisplatin alone in control of peritoneal carcinomatosis. The response rate was 5/7 (77%) for combined modality therapy vs. 2/9 (22%) for intraperitoneal chemotherapy alone. Both in vitro and in vivo studies suggest a role for interperitoneal therapy for control of refractory ascites in ovarian cancer.

Establishment and characterization of two new human ovarian cancer cell lines UWOV1 and UWOV2 and a subline UWOV2 (Sf) growing in serum-free conditions: growth characteristics, biochemical, and cytogenetic studies.

The establishment, growth, and characterization of two new continuously growing human ovarian cancer cell lines (UWOV1 and UWOV2) as well as a subline (UWOV2 Sf) grown in chemically defined, serum-free medium are described. The cell lines were derived from ascitic tumors of two patients suffering from cystadenocarcinomas of the ovary. Both UWOV1 and UWOV2 lines grow in anchorage-dependent fashion as monolayers, whereas UWOV2 (Sf) forms multilayered domelike structures. Cytogenetic studies revealed nonrandom abnormalities involving chromosomes 1 and 11 in all three cell lines. Secretion of soluble collagen was detected in all three lines. In addition, UWOV2 (Sf) produces and secretes large amounts of extracellular matrix material with an ordered fibrillar structure which may function as an attachment factor for the serum-free cells. These cell lines seem to be useful for further studies of the biology of human ovarian cancer.

Adriamycin cellular transport: methodological aspects.

A rapid and simple method for determination of cellular uptake of adriamycin is described. The method is based on the principle that active uptake is proportional to alterations of drug distribution, measured as a fraction of time, between suspending medium and cells, the volume of each having been accurately determined. Cellular drug uptake can be calculated by the use of a simple distribution formula. This method represents a compromise between indirect measurement of the loss of drug from suspending medium and direct measurement of drug uptake following cell separation, washing, and lysis. This method should be applicable to the measurement of cellular uptake of a wide range of drugs.

A comparison of three methods for titration of poliovirus vaccines.

Two methods for in vitro endpoint titration of poliovirus--the roller tube and the microtitration assay--were compared with each other and with the plaque assay, using secondary vervet monkey kidney cells and Vero cells as indicators. The roller tube method is the most reliable under difficult working conditions, but is otherwise cumbersome and expensive. The microtitre method is the most economical and the plaque assay the most sensitive. By suspending freshly trypsinized indicator cells with the virus dilutions before planting, it was possible to simplify the microtitre method considerably. The sensitivity of the plaque assay was improved for Vero cells by absorbing the virus onto freshly planted monolayers. The method was scaled down to a semi-micro level by using 24-well cell culture trays. The slower rate of plaque development under a low calcium overlay medium facilitated a more accurate plaque count.