Objective: To evaluate the age-related changes in active pumping in thoracic duct (TD) from 24-month-old Fisher-344 rats comparing with TD pumping in 9-month rats. Methods: Lymphatic diameters, contraction amplitude and frequency, ejection fraction, and fractional pump flow were determined in isolated TD preparations. Western blot analyses were performed to evaluate relative levels of eNOS and iNOS in 9- and 24-month-old TD. Results: Stretch-dependent regulation was altered in aged TD especially at higher levels of pressure: the negative inotropy, negative chronotropy and diminished minute pumping (2- to 3-fold decrease) were observed. Physiological NO/imposed-flow-dependent inhibition was completely abolished in aged TD, yet NO-synthase blockade by l-NAME (10-4 M) increased pumping in a flow-independent manner. Western blot analyses indicated that the relative levels of eNOS were decreased ∼ 7-fold in the 24-month-old TD when compared with 9-month-old TD; whereas iNOS levels were increased ∼ 10-fold in 24-month-old TD. Conclusions: These data provide the first evidence that stretch- and imposed-flow-dependent regulatory mechanisms are greatly altered in aged TD. These alterations of active pumping mechanisms in TD appear to be related with age-related disturbances in NO-dependent regulatory pathways, and may reflect diminished lymphatic muscle contractility as well as altered lymphatic endothelium function.

The lymphatic system is an extensive vascular network responsible for the transport of fluid, immune cells, proteins and lipids. It is composed of thin-walled vessels, valves, nodes and ducts, which work together to collect fluid, approximately 4 L/day, from the interstitium transporting it back to the systemic network via the great veins. The failure to transport lymph fluid results in a number of disorders and diseases. Lymphedema, for example, is a pathology characterized by the retention of fluid in limbs creating extreme discomfort, reduced mobility and impaired immunity. In general, there are two types of edema: primary edema, being those cases that are inherited (i.e. genetic predisposition), and secondary edema, which develop post-trauma or injury of the lymphatic vessels. With the onset of breast cancer and radiation therapies, the prevalence of secondary edema is on the rise. Clinical studies have shown that up to 80% of women who undergo nodal-dissection surgery develop lymphedema in their arms within 3-5 years of the surgery. Unfortunately, there is no cure or remedy for lymphedema stemming from our lack of understanding of the lymphatic system. The goal of this study was to evaluate lymph flow both experimentally and analytically to better understand the mechanisms regulating lymph transport. In particular we investigated the effects of pressure, volume loads and valve resistance on lymphatic function in the rat mesentery. Our experimental results were then used to develop computational and constitutive models to emulate the dynamic behavior of lymph transport. Collectively, the data illustrate the mechanics of lymphatic contractility and lymph flow. In particular, lymph flow and pumping significantly increased post edemagenic stress in the rat model. Furthermore, lymphangions exhibited highly nonlinear pressure-diameter responses at low pressures between 3-5 cmH2O. These experimental results strongly suggest the regulation of lymph flow via changes in pressure, shear stress and vessel diameter. Furthermore, the computational and constitutive models from this study provide great insight into lymphatic function characterizing the mechanical properties of a single pumping unit (i.e. lymphangion). These models will serve as valuable tools to further lymphatic research.

In patients affected by acute respiratory failure or acute respiratory distress syndrome (ARDS) the leading cause of death is failure of different vital organs other than the lungs, so called multiple organ dysfunction syndrome (MODS). The abdominal organs have a crucial role in the pathogenesis of this syndrome. There is a lack of knowledge regarding the mechanisms by which mechanical ventilation can affect the abdominal compartment. One hypothesis is that mechanical ventilation can interfere with abdominal fluid balance causing edema and inflammation. We addressed the question whether different levels of ventilatory support (mechanical ventilation with different levels of positive end-expiratory pressure, PEEP, and spontaneous breathing with or without PEEP) can influence abdominal edema and inflammation in both healthy and endotoxin-exposed animals. The effect on lymphatic drainage from the abdomen exerted by different degrees of ventilatory support was evaluated (paper I). We demonstrated that endotoxin increases abdominal lymph production, that PEEP and mechanical ventilation increase lymph production but also impede lymphatic drainage; spontaneous breathing improves lymphatic drainage from the abdomen. By adapting a non-invasive nuclear medicine imaging technique and validating it (paper II), we have been able to evaluate extravascular fluid accumulation (edema formation) in the abdomen over time (paper III) demonstrating that edema increases during endotoxemia, mimicking a sepsis-like condition, and that spontaneous breathing, compared to mechanical ventilation, reduces extravascular fluid. Pro-inflammatory cytokines TNF-α and IL-6 in intestinal biopsies are reduced during spontaneous breathing compared to mechanical ventilation. Abdominal edema results in increased intra-abdominal pressure (IAP): in paper IV we analyzed the effect of increased intra-abdominal pressure on the respiratory system. Pulmonary shunt fraction increased with high IAP both in healthy and LPS animals, resulting in decreased level of oxygenation. These changes are only partially reversible by reducing IAP. In conclusion, mechanical ventilation is a life-saving tool but the possible side effect at the extra-pulmonary level should be considered, and the introduction of some degree of spontaneous breathing when clinically possible is a suggested choice.