Tony J. Akl

Mesenteric lymph flow in adult and aged rats

The objective of study was to evaluate the aging-associated changes, contractile characteristics of mesenteric lymphatic vessels (MLV), and lymph flow in vivo in male 9- and 24-mo-old Fischer-344 rats. Lymphatic diameter, contraction amplitude, contraction frequency, and fractional pump flow, lymph flow velocity, wall shear stress, and minute active wall shear stress load were determined in MLV in vivo before and after N(ω)-nitro-L-arginine methyl ester hydrochloride (L-NAME) application at 100 μM. The active pumping of the aged rat MLV in vivo was found to be severely depleted, predominantly through the aging-associated decrease in lymphatic contractile frequency. Such changes correlate with enlargement of aged MLV, which experienced much lower minute active shear stress load than adult vessels. At the same time, pumping in aged MLV in vivo may be rapidly increased back to levels of adult vessels predominantly through the increase in contraction frequency induced by nitric oxide (NO) elimination. Findings support the idea that in aged tissues surrounding the aged MLV, the additional source of some yet unlinked lymphatic contraction-stimulatory metabolites is counterbalanced or blocked by NO release. The comparative analysis of the control data obtained from experiments with both adult and aged MLV in vivo and from isolated vessel-based studies clearly demonstrated that ex vivo isolated lymphatic vessels exhibit identical contractile characteristics to lymphatic vessels in vivo.

Lymph transport in rat mesenteric lymphatics experiencing edemagenic stress

To assess lymphatic flow adaptations to edema, we evaluated lymph transport function in rat mesenteric lymphatics under normal and increased fluid volume (edemagenic) conditions in situ.

Optofluidic phantom mimicking optical properties of porcine livers.

One strategy for assessing efficacy of a liver transplant is to monitor perfusion and oxygenation after transplantation. An implantable optical sensor is being developed to overcome inadequacies of current monitoring approaches. To facilitate sensor design while minimizing animal use, a polydimethylsiloxane (PDMS)-based liver phantom was developed to mimic the optical properties of porcine liver in the 630-1000 nm wavelength range and the anatomical geometry of liver parenchyma. Using soft lithography to construct microfluidic channels in pigmented elastomer enabled the 2D approximation of hexagonal liver lobules with 15mm sinusoidal channels, which will allow perfusion with blood-mimicking fluids to facilitate the development of the liver perfusion and oxygenation monitoring system.

Aging-related anatomical and biochemical changes in lymphatic collectors impair lymph transport, fluid homeostasis, and pathogen clearance

The role of lymphatic vessels is to transport fluid, soluble molecules, and immune cells to the draining lymph nodes. Here, we analyze how the aging process affects the functionality of the lymphatic collectors and the dynamics of lymph flow. Ultrastructural, biochemical, and proteomic analysis indicates a loss of matrix proteins, and smooth muscle cells in aged collectors resulting in a decrease in contraction frequency, systolic lymph flow velocity, and pumping activity, as measured in vivo in lymphatic collectors. Functionally, this impairment also translated into a reduced ability for in vivo bacterial transport as determined by time‐lapse microscopy. Ultrastructural and proteomic analysis also indicates a decrease in the thickness of the endothelial cell glycocalyx and loss of gap junction proteins in aged lymph collectors. Redox proteomic analysis mapped an aging‐related increase in the glycation and carboxylation of lymphatics endothelial cell and matrix proteins. Functionally, these modifications translate into apparent hyperpermeability of the lymphatics with pathogen escaping from the collectors into the surrounding tissue and a decreased ability to control tissue fluid homeostasis. Altogether, our data provide a mechanistic analysis of how the anatomical and biochemical changes, occurring in aged lymphatic vessels, compromise lymph flow, tissue fluid homeostasis, and pathogen transport.

Measuring contraction propagation and localizing pacemaker cells using high speed video microscopy

Previous studies have shown the ability of many lymphatic vessels to contract phasically to pump lymph. Every lymphangion can act like a heart with pacemaker sites that initiate the phasic contractions. The contractile wave propagates along the vessel to synchronize the contraction. However, determining the location of the pacemaker sites within these vessels has proven to be very difficult. A high speed video microscopy system with an automated algorithm to detect pacemaker location and calculate the propagation velocity, speed, duration, and frequency of the contractions is presented in this paper. Previous methods for determining the contractile wave propagation velocity manually were time consuming and subject to errors and potential bias. The presented algorithm is semiautomated giving objective results based on predefined criteria with the option of user intervention. The system was first tested on simulation images and then on images acquired from isolated microlymphatic mesenteric vessels. We recorded contraction propagation velocities around 10 mm/s with a shortening speed of 20.4 to 27.1 μm/s on average and a contraction frequency of 7.4 to 21.6 contractions/min. The simulation results showed that the algorithm has no systematic error when compared to manual tracking. The system was used to determine the pacemaker location with a precision of 28 μm when using a frame rate of 300 frames per second.

Optimizing probe design for an implantable perfusion and oxygenation sensor

In an effort to develop an implantable optical perfusion and oxygenation sensor, based on multiwavelength reflectance pulse oximetry, we investigate the effect of source–detector separation and other source-detector characteristics to optimize the sensor’s signal to background ratio using Monte Carlo (MC) based simulations and in vitro phantom studies. Separations in the range 0.45 to 1.25 mm were found to be optimal in the case of a point source. The numerical aperture (NA) of the source had no effect on the collected signal while the widening of the source spatial profile caused a shift in the optimal source-detector separation. Specifically, for a 4.5 mm flat beam and a 2.4 mm × 2.5 mm photodetector, the optimal performance was found to be when the source and detector are adjacent to each other. These modeling results were confirmed by data collected from in vitro experiments on a liver phantom perfused with dye solutions mimicking the absorption properties of hemoglobin for different oxygenation states.

Wireless Monitoring of Liver Hemodynamics In Vivo

Liver transplants have their highest technical failure rate in the first two weeks following surgery. Currently, there are limited devices for continuous, real-time monitoring of the graft. In this work, a three wavelengths system is presented that combines near-infrared spectroscopy and photoplethysmography with a processing method that can uniquely measure and separate the venous and arterial oxygen contributions. This strategy allows for the quantification of tissue oxygen consumption used to study hepatic metabolic activity and to relate it to tissue stress. The sensor is battery operated and communicates wirelessly with a data acquisition computer which provides the possibility of implantation provided sufficient miniaturization. In two in vivo porcine studies, the sensor tracked perfusion changes in hepatic tissue during vascular occlusions with a root mean square error (RMSE) of 0.135 mL/min/g of tissue. We show the possibility of using the pulsatile wave to measure the arterial oxygen saturation similar to pulse oximetry. The signal is also used to extract the venous oxygen saturation from the direct current (DC) levels. Arterial and venous oxygen saturation changes were measured with an RMSE of 2.19% and 1.39% respectively when no vascular occlusions were induced. This error increased to 2.82% and 3.83% when vascular occlusions were induced during hypoxia. These errors are similar to the resolution of a commercial oximetry catheter used as a reference. This work is the first realization of a wireless optical sensor for continuous monitoring of hepatic hemodynamics.

Intestinal perfusion monitoring using photoplethysmography

Abstract. In abdominal trauma patients, monitoring intestinal perfusion and oxygen consumption is essential during the resuscitation period. Photoplethysmography is an optical technique potentially capable of monitoring these changes in real time to provide the medical staff with a timely and quantitative measure of the adequacy of resuscitation. The challenges for using optical techniques in monitoring hemodynamics in intestinal tissue are discussed, and the solutions to these challenges are presented using a combination of Monte Carlo modeling and theoretical analysis of light propagation in tissue. In particular, it is shown that by using visible wavelengths (i.e., 470 and 525 nm), the perfusion signal is enhanced and the background contribution is decreased compared with using traditional near-infrared wavelengths leading to an order of magnitude enhancement in the signal-to-background ratio. It was further shown that, using the visible wavelengths, similar sensitivity to oxygenation changes could be obtained (over 50% compared with that of near-infrared wavelengths). This is mainly due to the increased contrast between tissue and blood in that spectral region and the confinement of the photons to the thickness of the small intestine. Moreover, the modeling results show that the source to detector separation should be limited to roughly 6 mm while using traditional near-infrared light, with a few centimeters source to detector separation leads to poor signal-to-background ratio. Finally, a visible wavelength system is tested in an in vivo porcine study, and the possibility of monitoring intestinal perfusion changes is showed.

Optimizing source detector separation for an implantable perfusion and oxygenation sensor

Each year thousands of patients are added to the waiting list for liver transplants. The first 7-10 days after transplant have proven to be the most critical in patient recovery and it is hypothesized that monitoring organ vital signals in this period can increase patient and graft survival rates. An implantable sensor to monitor the organ perfusion and oxygenation signals following surgery is being developed by our group. The sensor operates based on measuring diffuse reflection from three light emitting diodes (735, 805 and 940 nm). In this work the optimal source detector spacing to maximize oxygenation signal level is investigated for a portal vein model. Monte Carlo simulations provided signal levels and corresponding penetration depths as a function of separation between a point optical source and detector. The modeling results indicated a rapid decay in the optical signal with increasing distance. Through further analysis, it was found that there exists an optimal range of point source to detector spacing, between roughly 1 and 2 mm, in which the blood signal from the simulated portal vein was maximized. Overall, these results are being used to guide the placement and configuration of our probe for in vivo animal studies.

Fast Imaging System and Algorithm for Monitoring Microlymphatics

The lymphatic system is not well understood and tools to quantify aspects of its behavior are needed. A technique to monitor lymph velocity that can lead to flow, the main determinant of transport, in a near real time manner can be extremely valuable. We recently built a new system that measures lymph velocity, vessel diameter and contractions using optical microscopy digital imaging with a high speed camera (500fps) and a complex processing algorithm. The processing time for a typical data period was significantly reduced to less than 3 minutes in comparison to our previous system in which readings were available 30 minutes after the vessels were imaged. The processing was based on a correlation algorithm in the frequency domain, which, along with new triggering methods, reduced the processing and acquisition time significantly. In addition, the use of a new data filtering technique allowed us to acquire results from recordings that were irresolvable by the previous algorithm due to their high noise level. The algorithm was tested by measuring velocities and diameter changes in rat mesenteric micro-lymphatics. We recorded velocities of 0.25mm/s on average in vessels of diameter ranging from 54um to 140um with phasic contraction strengths of about 6 to 40%. In the future, this system will be used to monitor acute effects that are too fast for previous systems and will also increase the statistical power when dealing with chronic changes. Furthermore, we plan on expanding its functionality to measure the propagation of the contractile activity.