Effects of glucose supply on myeloma growth and metabolism in chemostat culture

The effects of the glucose supply on growth and metabolism of an SP2/0 derived recombinant myeloma cell line were studied in chemostat culture during growth on IMDM medium at a fixed dilution rate of 0.032 h^?1. Lowering of the feed medium glucose concentration from 25.0 to 1.4 mmol/L resulted in a decrease of steady-state viable cell concentration from 1.9 × 10⁹ L^?1, whereas viability remained above 90%. Mass balances indicated that only a minor amount of glucose was utilized via the TCA cycle irrespective of the glucose concentration in the feed medium. The apparent biosynthetic yield of cells from ATP was independent of the ratio between the specific glucose and glutamine consumption rate. It is concluded that the primary role of glucose is the provision of intermediates for anabolic reactions. In addition, glucose may play an indirect catabolic role in the process of glutaminolysis by providing the pyruvate for the transamination of glutamate to alanine and α-ketoglutarate. At low glucose concentrations in the feed medium, glutamine is probably the sole energy source for this myeloma in chemostat culture. © 1995 Wiley-Liss, Inc.     

Dielectric spectroscopy as a novel and convenient tool for the study of the shear sensitivity of plant cells in suspension culture.

Plant cell suspensions of different species and different age were subjected to hydrodynamic stress while following the decline in the volume fraction of intact cells by measuring the permittivity of the cell suspension at radio frequencies. Results were compared with the fresh weight, dry weight, packed cell volume and cell number of the suspensions. At first a rapid decline is seen as the most shear-sensitive cells are broken up, followed by a slower decline as less sensitive cells are broken up. The sensitivity of the cells to shear stress depended strongly on the cell line used but only slightly on their age, older cells being more sensitive. The dependence of the shear sensitivity on the cell line might be an effect of the species investigated, the culturing conditions of the cell line, or both. It was found that cells that grow in a finely dispersed suspension are much less prone to shear stress than is often assumed.     

Evaluation of enzyme histochemical observations for metabolic studies. A combined histochemical and biochemical investigation of experimental induced muscle-changes

Summary The degree of conformity between enzyme histochemical (part I) and biochemical observations was studied in a variable model. Rabbit skeletal muscle altered by reinnervation was used. The most important feature of the biochemical part of this investigation is the constancy of the activity ratio of the enzymes phosphofructokinase (PFK), which is ratelimiting for the glycolysis and l-glycerol-3-phosphate: acceptor oxidoreductase (GPOX) in spite of the marked metabolic changes after cross-reinnervation. In consequence it is most likely that in case of pathologically altered tissue one may also rely on the histochemical GPOX-reaction as an indication of the PFK-activity, which cannot be demonstrated histochemically to any reliable degree.In order to evaluate the indicator significance of the GPOX-reaction, the histochemical estimations (part I) were compared with the biochemical results. This comparison disclosed a striking parallellism per type of muscle in discerning the presence or the absence of metabolic changes after cross-reinnervation or auto-reinnervation. Quantitatively though there were some variations especially for the m. soleus. Consequently the histochemical impressions of the investigated enzymes are applicable in predicting the biochemical results within this model, but cannot be used as a substitute. It was concluded that where only small amounts of tissue are available histochemical examination offers a reliable screening method for muscle diseases and can indicate a more appropriate biochemical investigation to confirm the diagnosis.This study was taken from the Ph. D. thesis of A. C. Jöbsis (1971).     

Detecting emerging transmissibility of avian influenza virus in human households

Accumulating infections of highly pathogenic H5N1 avian influenza in humans underlines the need to track the ability of these viruses to spread among humans. A human-transmissible avian influenza virus is expected to cause clusters of infections in humans living in close contact. Therefore, epidemiological analysis of infection clusters in human households is of key importance. Infection clusters may arise from transmission events from (i) the animal reservoir, (ii) humans who were infected by animals (primary human-to-human transmission), or (iii) humans who were infected by humans (secondary human-to-human transmission). Here we propose a method of analysing household infection data to detect changes in the transmissibility of avian influenza viruses in humans at an early stage. The method is applied to an outbreak of H7N7 avian influenza virus in The Netherlands that was the cause of more than 30 human-to-human transmission events. The analyses indicate that secondary human-to-human transmission is plausible for the Dutch household infection data. Based on the estimates of the within-household transmission parameters, we evaluate the effectiveness of antiviral prophylaxis, and conclude that it is unlikely that all household infections can be prevented with current antiviral drugs. We discuss the applicability of our method for the detection of emerging human-to-human transmission of avian influenza viruses in particular, and for the analysis of within-household infection data in general.     

Ultraviolet light provides a major input to non-image-forming light detection in mice

The change in irradiance at dawn and dusk provides the primary cue for the entrainment of the mammalian circadian pacemaker. Irradiance detection has been ascribed largely to melanopsin-based phototransduction [1-5]. Here we examine the role of ultraviolet-sensitive (UVS) cones in the modulation of circadian behavior, sleep, and suprachiasmatic nucleus (SCN) electrical activity. UV light exposure leads to phase-shifting responses comparable to those of white light. Moreover, UV light exposure induces sleep in wild-type and melanopsin-deficient (Opn4(-/-)) mice with equal efficacy. Electrical recordings from the SCN of wild-type mice show that UV light elicits irradiance-dependent sustained responses that are similar to those induced by white light, with characteristic fast transient components occurring at the light transitions. These responses are retained in Opn4(-/-) mice and preserved under saturating photopic conditions. The sensitivity of phase-shifting responses to UV light is unaffected by the loss of rods but is severely attenuated by the additional loss of cones. Our data show that UVS cones play an important role in circadian and sleep regulation in mice.     

Human liver slices as an in vitro model to study toxicity-induced hepatic stellate cell activation in a multicellular milieu

INTRODUCTION: Hepatic stellate cell (HSC) activation is a key event in wound healing as well as in fibrosis development in the liver. Previously we developed a technique to induce HSC activation in slices from rat liver. Although this model provides a physiologic, multicellular milieu that is not present in current in vitro models it might still be of limited predictive value for the human situation due to species-differences. Therefore, we now aimed to evaluate the applicability of human liver slices for the study of HSC activation. METHOD: Liver slices (8 mm diameter, 250 microm thickness) were generated from human liver tissue and incubated for 3 or 16 h with 0-15 microl of carbon tetrachloride (CCl4) after which ATP-content and expression levels of HSC (activation) markers was determined. RESULTS: Human liver slices remained viable during incubation as shown by constant ATP levels. Incubation with CCl(4) caused a dose-dependent decrease in viability and an increase in mRNA expression of the early HSC activation markers HSP47 and alphaB-crystallin, but not the late markers for HSC activation, alphaSMA and pro-collagen 1a1. Synaptophysin mRNA expression remained constant during incubation with or without CCl4, indicating a constant number of HSC in the liver slices. CONCLUSION: We developed a technique to induce early toxicity-induced HSC activation in human liver slices. This in vitro model provides a multicellular, physiologic milieu to study mechanisms underlying toxicity-induced HSC activation in human liver tissue.     

(R)-roscovitine (CYC202, Seliciclib) sensitizes SH-SY5Y neuroblastoma cells to nutlin-3-induced apoptosis

In this study, we have analyzed the consequences, on several neuroblastoma cell lines, of combined treatments with (R)-roscovitine (CYC202, Seliciclib), a CDK inhibitory drug, and nutlin-3, a p53 activating drug. Both compounds were found to synergize, causing significant levels of apoptosis in cultured cells when combined at sublethal concentrations. In SH-SY5Y cells, Bcl-XL protein overexpression protected from apoptosis induced by either nutlin-3 alone or the (R)-roscovitine plus nutlin-3 association but failed to prevent apoptosis triggered by (R)-roscovitine alone. Moreover, Western blot studies showed that (R)-roscovitine increased nutlin-3-mediated p53 stabilization. Therefore, we conclude the contribution of (R)-roscovitine to the synergism is basically the sensitization of SH-SY5Y cells to the action of nutlin-3 on p53. The relevance of this pharmacological synergism with respect to the treatment of neuroblastoma is discussed.     

Perioperative IFN-alpha to avoid surgically induced immune suppression in colorectal cancer patients

Surgical treatment of colorectal cancer is associated with postoperative immunosuppression, which might facilitate dissemination of tumor cells and outgrowth of minimal residual disease/(micro) metastases. Minimal residual disease has been shown to be of prognostic relevance in colorectal cancer. Therefore, stimulation of (anti-tumor) immune responses may be beneficial in the prevention of metastases formation. Important anti-tumor effector cells, which serve this function, are natural killer (NK) cells, CD8+ lymphocytes (CTL), dendritic cells (DC) and macrophages. In this review the immunomodulating properties of IFN-alpha are discussed, with a particular focus on perioperative stimulation of immune function in cancer patients. IFN-alpha is known to enhance innate immune functions such as stimulation of NK cells, transition from innate to adaptive responses (activation of DC) and regulating of CD8+ CTL activity and memory. Moreover, it exerts direct antitumor effects by regulating apoptosis and cell cycle. In several clinical trials, perioperative administration of IFN-alpha has indeed been shown to improve T cell responsiveness, prevent impairment of NK cell cytotoxicity and increase expression of activation markers on NK, T and NKT cells. In a clinical pilot study we showed in colorectal cancer patients that received perioperative IFN-alpha enhanced activation markers on T cells and NK cells, combined with better-preserved T cell function as indicated by phytohemaggluttinin skin tests. In the liver of these patients significantly more CD8+ T cells were found. In conclusion, IFN-alpha provides an effective adjuvant in several forms of cancer and improves several postoperative immune functions in perioperative administration. However, larger clinical trials are necessary to investigate effects on disease-free and overall survival.