Weightlessness impacts the cardiovascular system, altering the function and structure of the body’s arterial circulation. Dr. Michael D. Delp is studying animals exposed to space flight-like conditions to examine the molecular mechanisms involved in these changes. He will also evaluate alterations to lymphatic function, which impact body fluid distribution.
Circulatory Remodeling with Simulated Microgravity
Michael D. Delp, Ph.D.
Texas A&M University
The proposed studies were designed address the effects of simulated microgravity on the arterial and lymphatic portions of the circulation. Using the hindlimb unloaded rat as a ground-based animal model, the general aim of this proposal is to determine the effects of simulated microgravity on 1) the molecular mechanisms mediating structural remodeling of the arterial resistance vasculature, and 2) the ability of the lymphatics to generate and modulate lymph flow.
Specific Aim 1:
To identify early regulatory events leading to hypertrophic remodeling of cerebral arteries in response to hindlimb unloading. We will characterize indicators of cell proliferation to determine whether increased medial thickness previously reported is due to cell growth. Utilizing RT-PCR, in situ hybridization and immunohistochemistry, we will identify key signaling mediators that are involved in this process. Our initial studies will concentrate on nitric oxide (NO) and endothelin (ET), both of which have been proposed to mediate mechanically induced vascular remodeling in other systems. Key cell growth, survival and differentiation markers of vascular smooth muscle cells will be examined to better determine their role in hypertrophic remodeling of the cerebral arteries.
Specific Aim 2:
To characterize signaling events leading to atrophy of resistance arteries in the soleus and gastrocnemius muscle in response to hindlimb unloading. Resistance arteries in the soleus muscle have been shown to atrophy in response to hindlimb unloading in a manner that leads to decreases in luminal diameter as a result of circumferential atrophy of smooth muscle cells (i.e., a decrease in muscle length). Intraluminal shear stress has been shown to decrease during the initial unloading period, but as remodeling occurs, shear stress returns to control levels. The goal of this aim will be to determine if there is increased susceptibility to apoptosis in smooth muscle cells with reduced shear, to evaluate nitric oxide synthase (NOS) activity and expression, and to determine if activation of matrix metalloproteinase activity is up regulated and contributes to decreased luminal diameter. In contrast to the apparent shear stress-mediated remodeling of soleus muscle resistance arteries, reduced transmural pressure appears to be the primary stimulus for remodeling of resistance arteries from the gastrocnemius muscle. The hindlimb unloading-induced remodeling of the gastrocnemius muscle resistance arteries does not involve alterations in the vessel diameter, but rather consists of a decrease in media thickness that appears to occur as a result of radial atrophy of smooth muscle cells (i.e., decreased thickness of smooth muscle cells). We will utilize cellular markers of apoptosis to quantify susceptibility to cell death. RT-PCR, in situ hybridization and immunohistochemistry will be used to quantify expression of growth factors, survival factors and contractile proteins in these vessels in response to hindlimb unloading.
Specific Aim 3:
To evaluate the effects of hindlimb unloading on the ability of the lymphatics from different regions of the body to generate and modulate lymph flow, and thus, regulate overall body fluid homeostasis. It has been demonstrated that acute change in the transmural pressure, luminal flow and outflow resistance will modulate lymph contractile activity and lymph flow. Furthermore, it is known that the special contractile characteristics of the lymphatics are reflected in the expression of contractile proteins within the lymphatic muscle cells. Given that hindlimb unloading induces tissue fluid shifts from the lower to the central and upper portions of the body, it is likely that these tissue fluid shifts will differentially alter the normal physical environment of the lymphatics in different lymphatic drainages. Thus, similar to that found in the arterial circulation, we hypothesize that chronic changes in the normal physical environment of the lymphatics (i.e., changes in tissue fluid pressure, lymph pressure and central venous pressure) will alter both the lymphatic contractile function and the expression of contractile proteins within the lymphatic muscle. Specifically, we will evaluate lymphatic contractile function and contractile protein expression from five regions of the body where significant fluid shifts are known to occur in response to microgravity, as well as in tissues where the majority of the bodys lymph is produced.
The present proposal is synergistically related to several projects within the cardiovascular team:
- The current project PI (Michael Delp) will assist the PI of another NSBRI cardiovascular project, Dr. Vince Cassone, with studies to determine the effects of hindlimb unloading on circadian changes in cardiac output and blood flow distribution;
- The current studies determining the effects of simulated microgravity on the arterial and lymphatic portions of the circulation are complimentary to those of Drs. Artin Shoukas and Dan Berkowitz, who will investigate the effects of hindlimb unloading on the venous portion of the circulation. Thus, these projects will provide a comprehensive investigation of the peripheral circulation in hindlimb unloaded rats, and;
- A collaboration between the current project PI (Michael Delp) and the PI of another NSBRI cardiovascular project, Dr. Chester Ray, was established to determine the effects of microgravity and hindlimb unloading on cardiac mass in rats. This project was funded, in part, by both of the current NSBRI grants to these PIs, and the results have been recently published.
Research Program Accomplishments
The initial studies were designed to determine the mRNA and protein expression of endothelial nitric oxide synthase (eNOS) in cerebral and skeletal muscle resistance arteries, which has been proposed to mediate mechanically induced vascular remodeling in other systems (Specific Aims 1 and 2). In the cerebral arteries, eNOS mRNA expression was not different between control and hindlimb unloaded rats after 1, 14 and 28 days of hindlimb unloading. However, eNOS protein levels are significantly depressed in the middle cerebral artery (MCA) after 14 days of hindlimb unloading.
The current results indicate that eNOS may be directly involved in the remodeling of the cerebral vascular hypertrophy induced by hindlimb unloading in rats. These results were presented at the Humans in Space Symposium, and a manuscript is in preparation.
Lymphatic contractile function from the mesenteric lymphatics and thoracic ducts have been tested and characterized from control rats, and the results have been published. Furthermore, the effect of hindlimb unloading has been determined to diminish the contractile function in these lymphatic vessels. More specifically, there is a 50-75 percent reduction in resting tone of lymphatic vessels, a 30-60 percent reduction in phasic contraction frequency of the lymph pump, a 60-80 percent reduction in the strength of phasic contractions of the lymph pump, and a significant reduction in the pressure-sensitive stimulation of the lymph pump. These results of simulated microgravity have been presented at the Bioastronautics Meeting and the Humans in Space Symposium. A manuscript also is in preparation.
One question in the cardiovascular area has been whether microgravity induces cardiac atrophy. We recently reported that results from rats flown for one week on the Spacelab 3 mission demonstrate that cardiac atrophy does not occur with short-term exposure to microgravity in rats (Ray et al. J Appl Physiol 91: 1207-1213, 2001). Similarly, we found that neither one week nor four weeks of hindlimb unloading induced cardiac atrophy or altered the peak rate of rise in left ventricular pressure, and index of myocardial contractility (Ray et al. J Appl Physiol 91: 1207-1213, 2001). However, there are studies in the literature reporting cardiac atrophy in hindlimb unloaded rats. To determine whether the cardiac atrophy reported in the literature may be related to caloric deficits in some hindlimb unloaded rats, we plotted heart mass as a function of body mass from all studies reporting these variables in the literature. We found that in all cases where cardiac atrophy was reported, there was a substantial corresponding loss of body mass. Therefore, these findings indicate that cardiac atrophy and dysfunction are not adverse consequences of short-term microgravity or long-term simulations of microgravity when body mass is fairly well maintained.