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This review deals with the deleterious effects of cigarette smroking on the microcirculation, both in terms of morphological (i.e., vessel wall injury, capillary loss) and functional aspects. The latter concerns predominantly changes in tissue perfusion and its regullatory mechanisms (i.e., reactive hyperemia, sequestration of blood cells in the microcirculation). The mechanisms of action of cigarette smoking on the microcirculation include compromised endothelial-dependent voasorelaxation, platelet aggregation, emdothelial cell dysfunction and the activation of circulating leukocytes. Through these mechanisms, cigarette smoking elicits the aggregation and adhesion of leukocytes and/or platelets to the microvascular endothelium in venules and arterioles, as assessed by intravital fluorescence microscopy in the hamster skinfold chamber model. This model has allowed us to learn more about the participation of reactive oxygen species, inflammatory mediators, and adhesion molecules in the orchestration of microcirculatory dysfunction after cigarette smoking. In the final part of this review, the clinical consequences of microcirculatory dysfunction are discussed and an outlook is offered on potential prophylactic interventions (i.e., antioxidant vitamins) aimed at abrogating the deleterious action of cigarette smoking on the microcirculation.

Background: Study end-points in microcirculation research are usually video-taped images rather than numeric computer print-outs. Analysis of these video-taped images for the quantification of microcirculatory parameters usually requires computer-based image analysis systems. Most software programs for image analysis are custom-made, expensive, and limited in their applicability to selected parameters and study end-points., Methods and Result: We demonstrate herein that an inexpensive, commercially available computer software (Adobe Photoshop), run on a Macintosh G3 computer with inbuilt graphic capture board provides versatile, easy to use tools for the quantification of digitized video images. Using images obtained by intravital fluorescence microscopy from the pre- and postischemic muscle microcirculation in the skinfold chamber model in hamsters, Photoshop allows simple and rapid quantification (i) of microvessel diameters, (ii) of the functional capillary density and (iii) of postischemic leakage of FITC-labeled high molecular weight dextran from postcapillary venules. We present evidence of the technical accuracy of the software tools and of a high degree of interobserver reliability., Conclusion: Inexpensive commercially available imaging programs (i.e., Adobe Photoshop) provide versatile tools for image analysis with a wide range of potential applications in microcirculation research.

Sepsis is a frequent complication of multiple organ dysfunction syndrome and remains a major problem of intensive care medicine. It is also a common factor in the final cause of death in hospital populations. Clinical observations, assisted by invasive monitoring techniques as well as pathological-anatomical studies, clearly indicate that microcirculatory dysfunction lies at the centre of sepsis pathogenesis. Numerous animal models, from rodents to primates, many of which employ bacteria or their toxins, especially endotoxins, have helped to shed light on the pathomechanisms leading to this dysregulation in the peripheral circulation. Among these are activation of humoral and cellular inflammatory mediator systems, with special emphasis on neutrophil-endothelial interactions, affecting endothelial barrier function and vasoregulation and ultimately leading to severely perturbed oxygen transport and utilization. In vitro studies have provided more insight into the molecular mechanisms involved in this microcirculatory dysfunction, although much more attention must be directed towards microvascular endothelial cells and the role of heterogeneity of response in various vascular beds. These experimental data must in turn be validated by comparing with the human in situ situation, both clinical and morphological. This review aims at a critical appraisal of the clinical and experimental evidence for sepsis-induced dysregulation of the microcirculation and how knowledge of the underlying cellular and molecular pathology could be used to make therapy more rational and effective. To date, therapeutic approaches, such as anti-cytokine and anti-oxidant regimens, which have been highly successful in experimental models, have failed to demonstrate clinical efficacy. Newer approaches, such as targeting the coagulation system, nitric oxide synthesis or intracellular signal transduction, are also discussed. The necessity to focus on the role of anti-inflammatory mediators, as well as the pathogenetic significance of important molecular groups, such as the heat shock proteins, which until now have been given scant attention, will be stressed., (Copyright 2000 John Wiley & Sons, Ltd.)

Aimed at improving animal fertility and health, diets for farm and laboratory animals have over the last few years been supplemented with increasing amounts of the antioxidant vitamin E. We now demonstrate by intravital microscopy that feeding hamsters with a vitamin E-supplemented "standard" rodent diet (60 ppm vitamin E) significantly reduces the microvascular manifestations of ischemia/reperfusion injury when compared to animals fed a nonsupplemented diet. Postischemic leukocyte adhesion to venular endothelium was reduced from 770 +/- 204 cells/mm2 at 24 h after reperfusion in control animals on the nonsupplemented diet to 403 +/- 105 cells/mm2 in animals on the "standard" rodent diet (means +/- SD, n = 7 animals per group, p < 0.01). Animals on the nonsupplemented diet showed a dramatic loss of capillary perfusion density until 7 days after reperfusion (to 21 +/- 13% of preischemic baseline values), whereas this loss was significantly attenuated (to 71 +/- 12% of preischemic values, p < 0.01) in animals on the "standard" rodent diet. No difference in the extent of reperfusion injury was seen between animals on the "standard" rodent diet and animals on diets with substantially higher vitamin E supplements (300 ppm-30,000 ppm). Besides underscoring the benefit of vitamin E in reducing the extent of ischemia/reperfusion injury, this study raises the concern that vitamin E supplements in "standard" laboratory animal diets may have a far-reaching impact on biomedical research by jeopardizing established animal models of disease.

As observations in vitro become more sophisticated, it is increasingly important to be able to assess their relevance to the intact animal. Intravital microscopy provides an advanced set of tools for the dissection of the in vivo situation.

Reperfusion after transient tissue ischemia constitutes an irrevocable need to preserve tissue viability. However, release of prolonged ischemia will either result in failure of the microcirculation to reperfusion (no-reflow) and thus the prolongation of hypoxia, or in restoration of blood flow resulting in reoxygenation of the inflicted tissue. While ischemia damages the tissue primarily through hypoxia-induced depletion of energy stores, reoxygenation paradoxically contributes to tissue damage through the formation of oxygen radicals, the release of chemoattractant mediators (TNF, IL-1, LTB4), and the activation of circulating polymorphonuclear leukocytes (PMNs). Through the action of chemoattractant mediators and the upregulation of leukocytic (CD11/CD18) and endothelial adhesion receptors (ICAM, GMP-140), activated PMNs adhere to the endothelium, release further chemoattractants and oxygen radicals and undertain a vicious circle, which will ultimately result in further tissue damage. Both the no-reflow phenomenon and the events initiated by reflow--termed herein as the reflow-paradox--contribute to the failure of the nutritive microvascular perfusion and loss of tissue viability following ischemia and reperfusion.

Epidemiologic observations and experimental studies have demonstrated a protective effect of dietary fish oil on the clinical manifestations of ischemia-reperfusion injury. To investigate the underlying mechanisms, we used the dorsal skinfold chamber model for intravital fluorescence microscopy of the microcirculation in striated muscle of awake hamsters. In control hamsters (n = 7), reperfusion after a 4-hr pressure-induced ischemia to the muscle tissue elicited the adhesion of fluorescently stained leukocytes to the endothelium of postcapillary venules, capillary obstruction, and the break-down of endothelial integrity. These microvascular manifestations of ischemia-reperfusion injury were significantly attenuated in animals (n = 7) when fed with a fish oil-enriched diet for 4 weeks prior to the experiments. In leukocyte total lipids, the fish oil diet resulted in a substantial displacement of arachidonic acid, the precursor of the potent adhesion-promoting leukotriene (LT) B4, by fish oil-derived eicosapentaenoic acid, the precursor of biologically less potent LTB5, emphasizing the mediator role of LTB4 in ischemia-reperfusion injury. These results suggest that the preservation of microvascular perfusion by dietary fish oil contributes to its protective effects on the clinical manifestations of ischemia-reperfusion injury.