Geometric Resistance and Microvascular Network Architecture of Human Colorectal Carcinoma

Objective : To measure the geometric resistance to blood flow in human colorectal carcinoma. Although tumor blood flow is of central importance in both the detection and the treatment of cancer, the determinants of blood flow through the neoplastic circulation are poorly understood. Methods : Human colorectal carcinomas (tissue weight = 272 g ± 43 g (SD), n = 6) were perfused ex vivo with a buffered physiological salt solution of known viscosity at flow rates ranging from 2.5 to 40 ml/min and perfusion pressures from 8 to 100 mm Hg. The geometric resistance was determined from the slope of the pressure-flow curve. For examination of the principal determinant of geometric resistance, the vascular architecture, one of the tumors was perfused with Batson's No. 17 polymer and macerated in KOH to produce a positive vascular cast that was used for measurement of vascular branching patterns and dimensions. Results : The pressure-flow relationship was linear at perfusion pressures above 40 mm Hg, and the geometric resistance, z₀, was constant at approximately 6.5 ± 10⁹g/cm³. Below 40 mm Hg, z₀ increased rapidly. The architecture of the arteriolar and capillary networks of human colorectal carcinoma is similar to those of experimental rodent tumors. Capillaries in planar and nonplanar mesh-works had mean segment diameters of 11 ± 2 and 9.6 ± 2 μm, lengths of 46 ± 24 and 107 ± 40 μm, and intercapillary distances of 46 ± 13 and 74 ± 24 μm, respectively. Conclusions : The geometric flow resistance in neoplastic tissue is 1–2 orders of magnitude higher than that observed in normal tissues. A decrease in functional vascular cross-sectional area may explain the additional increase in resistance at small perfusion pressures. The observed flow resistance may be due to the specialized arteriolar and capillary network architecture, pressures exerted by proliferating cancer cells, and/or coupling between vascular and extravascular flow. These observations demonstrate that tumor vascularity alone may not be indicative of flow resistance or tumor susceptibility to blood-borne therapeutic agents.     

Assessment of the potential in vivo genotoxicity of fluoranthene

Fluoranthene is a ubiquitous environmental pollutant. Althoughfluoranthene is mutagenic in bacterial and mammalian in vitrocell systems following metabolic activation by rat liver fraction,information on in vivo mutagenicity is lacking and studies ontumour initiating activity in mice are equivocal. In the presentstudy, the potential genetic hazard to man was assessed usingthe mouse bone marrow micronucleus and rat liver unscheduledDNA synthesis test systems. Fluoranthene did not show any evidenceof genotoxicity in either of the in vivo assays following acuteoral administration at levels of up to 2000 mg/kg b.w. ¹To whom correspondence should be addressed     

The regional vulnerability to hypoglycemia-induced neurotoxicity in organotypic hippocampal culture: protection by early tetrodotoxin or delayed MK-801.

Profound hypoglycemia selectively damages CA1 and the dentate gyrus of the hippocampus. We have examined the time course of hippocampal neuronal injury in organotypic cultures following in vitro "hypoglycemia," using the fluorescent vital dye propidium iodide to observe directly the regional distribution of early neuronal membrane injury in living cultures. The in vivo hippocampal pattern of hypoglycemic injury was reproduced by a 2 hr exposure to glucose-free media, which resulted in simultaneous, selective propidium staining of CA1 and the dentate gyrus starting by 4 hr after exposure. After 24 hr of recovery, CA3 remained spared. A similar pattern of propidium staining was produced by incubation of cultures for briefer periods in glucose-free medium containing 5 mM 2-deoxyglucose (2-DG) to inhibit glycolysis. This "hypoglycemic" pattern and time course of neuronal injury was mimicked by 300 microM aspartate but not by glutamate. The NMDA receptor antagonists MK-801 and CPP, but not the relatively selective non-NMDA receptor antagonist 6-cyano-7-dinitroquinoxaline-2,3-dione, prevented the development of propidium staining. MK-801 protected against injury even if added to the recovery media 30 min after the insult, while TTX (10 microM) protected only if added by the end of the exposure. The appearance of propidium staining after 4-6 hr of recovery was well correlated with histological observation of pyknotic neuronal nuclei in the injured regions. The characteristic hippocampal regional vulnerability of CA1 and the dentate gyrus to injury following profound hypoglycemia can be reproduced in organotypic hippocampal culture and appears to be mediated both by an early TTX-sensitive component and by a more prolonged period of toxic NMDA receptor activation, extending for at least 30 min into the recovery period.     

Preclinical pharmacologic studies of the new antitumor agent carmethizole (NSC-602668) in the mouse and beagle dog

Summary The chemical breakdown of carmethizole [1-methyl-2-methylthio-4,5-bis-(hydroxymethyl)imidazole-4,5-bis(N-methylcarbamate)hydrochloride] and its pharmacokinetics in the mouse and beagle dog were studied. Carmethizole was relatively unstable in aqueous media, having a half-life of 1 h in 0.9% sodium chloride, human whole blood, human plasma, and dog urine at 37°C. Its major breakdown product in 0.9% sodium chloride and pH 5.0 sodium phosphate buffer was carmethizole diol. When carmethizole was added to pH 7.0 or pH 9.0 sodium phosphate buffer, the major breakdown product was carmethizole diol-4-monophosphate. Carmethizole reacted directly with glutathione at pH 8.0, forming a glutathione adduct of carmethizole monocarbamate. Elimination of the drug from the plasma of the beagle dog following i.v. bolus doses of 22.4 and 4.3 mg/kg was biphasic. At these doses the terminal half-life was 39 and 46 min, respectively, and the respective total body clearance was 4.6 and 7.7 ml/min per kg. The 22.4 mg/kg dose was lethal to the beagle dog by day 4. Elimination of carmethizole from the plasma of mice following an i. v. bolus dose of 115 mg/kg was monoexponential, with a half-life of 11.6 min and a total body plasma clearance of 43.6 ml/min per kg. When the drug was infused at 230 mg/kg over 8 h into mice, the total body clearance was 40.8 ml/min per kg. Following the i.v. bolus administration of carmethizole to mice, 30% of the total dose was excreted in urine over 3 h as carmethizole diol, 10%, as carmethizole diol-sulfate, 3.4%, as carmethizole 4-monocarbamate, and 2.4%, as unchanged drug.     

Real-time nested multiplex PCR for the detection of herpes simplex virus types 1 and 2 and varicella zoster virus

One hundred forty-nine specimens were tested in a LightCycler nested multiplex polymerase chain reaction (LCnmPCR) for Herpes simplex virus (HSV)1, HSV2, and VZV. Eighty-one were from genitourinary medicine (GUM) patients and the other 68 specimens were from other patients with skin lesions. The results were compared to a conventional multiplex nested PCR (nmPCR) using agarose gel electrophoresis. Twenty-five specimens were positive in both assays for HSV1 and 29 were positive for VZV. For HSV2 there were 27 positive in the LCnmPCR and 26 positive in the nmPCR assay. The melting temperatures (Tms) of each target were different with a mean of 84.75 degrees C for HSV1, 88.57 degrees C for HSV2, and 83.62 degrees C for VZV. The melting curves of positive specimens directly overlaid the melting curves of the positive controls in the assay. The LCnmPCR assay is a convenient alternative to conventional PCR using agarose gel electrophoresis. It improves specimen turnaround time by eliminating the need for gel electrophoresis, transillumination, and gel photography. It also shows increased sensitivity for HSV2 over our standard assay. This LCnmPCR reduces further the possibility of amplicon contamination with nested PCR protocols.     

Leukocyte apoptosis and its significance in sepsis and shock

Sepsis and multiple organ failure continue to be significant problems among trauma, burn, and the critically ill patient population. Thus, a number of laboratories have focused on understanding the role of altered apoptotic cell death in contributing to immune and organ dysfunction seen in sepsis and shock. Immune cells that undergo altered apoptotic changes include neutrophils, macrophages, dendritic cells, as well as various lymphocyte populations. Evidence of epithelial as well as endothelial cell apoptotic changes has also been reported. Although mediators such as steroids, tumor necrosis factor, nitric oxide, C5a, and Fas ligand (FasL) appear to contribute to the apoptotic changes, their effects are tissue- and cell population-selective. As inhibiting Fas-FasL signaling (e.g., gene deficiency, Fas fusion protein, or Fas short interfering RNA administration), caspase inhibition (caspase mimetic peptides), and/or the overexpression of downstream antiapoptotic molecules (e.g., Bcl-2, Akt) improve survival of septic mice, it not only demonstrates the pathological significance of this process but points to novel targets for the treatment of sepsis.