This experiment confirms that LFVP (50 Hz, ~4.0 mm), periodically applied during the diastolic phase of a series of arterial like pressure pulses and engaged upon a 4 cm meat barrier provides enhanced clot disruptive effects in an underlying coronary like tube system. This is a first demonstration that LFVP engaged across a chest wall sized barrier can penetrate to yield potential thrombo-clearing effects, with the effects accentuated in conjunction with a remotely delivered thrombolytic agent.
The first notable finding in our experiment was an obvious development of clot length fluid channels in all LFVP samples (i.e. SK enriched and non-enriched) which were completely absent in all control samples. Such channeling alongside or within a clot would certainly predict a therapeutic benefit as early reflow along with enhanced penetration of systemically delivered therapeutic drug agents would undoubtedly be promoted. As LFVP is known to produce convective currents[33, 34], it is probable that LFVP induced shear forces within the HS (+/− SK) solution likely acted upon (or eroded) the loosely aggregated platelet surface along the young clot’s edges. It is also possible that vibro-agitation with accompanying catheter wall deformations may have added to the erosive effect, and at least partly caused the clot to unfurl, or lay more extended lengthwise along the catheter’s lumen (which was a general observation noted in our experiment). Channel development generally occurred faster and more extensively in the SK enriched vs. non-enriched group.
A second notable finding was that while we observed a consistent but somewhat “under-whelming” improvement in Percent Clot Dissolution in the non-SK enriched LFVP group versus control (i.e. 21% - which might be to slight a change to predict a clinical benefit), we however more than doubled this improvement (to 48%) in the SK enriched group. The improved effectiveness of LFVP with remotely administered SK (a pairing which we have dubbed “Vibrinolytic Therapy”) is likely explained by a combination of introduced proximal fluid turbulent mixing (as a means for enhanced thrombolytic delivery - discussed more thoroughly below), along with the above mentioned development of clot length fluid channels which would enhance clot surface area exposure and fibrin binding sites to the penetrating SK molecule.
A third notable finding was an observed marked enhancement of dissolved clot constituent mixing in the LFVP samples (as evidenced by the diffusive permeation of partially transparent red color within the full height of the catheter segment and up into the connecting line), which was substantially unobserved in the control samples. This finding, besides confirming that significant clot dissolution had taken place, supports our suspicions that LFVP may have promoted delivery of SK towards the clot interface by a turbulent mixing phenomenon. Indeed, the relatively slow mass transport of systemically introduced clot busters down occluded thrombosed arteries and into clots (where the process is by in large dictated by arterial based filtration pressure with non-facilitated diffusion) has certainly been an Achilles Heel in the mechanistic effectiveness of conventional, passively introduced IV thrombolytic therapy. Clearly that remotely delivered SK in our experiment was shown to be negligibly effective in clot dissolution without LFVP and note-ably effective with LFVP supports the view that a degree of enhanced drug transport had most probably taken place.
That LFVP enhances mixing between two adjacent fluids is supported in fluid mechanics. It has been solidly established that mix ability of solutes and parallel fluids within an agitated fluid system is several orders of magnitude greater than what is seen in laminar (and especially absent) flow, due to the introduced random velocity and density gradients which cause eddies and vortices which greatly accelerate diffusion and mass transport[36, 37]. Indeed the correlation of increased mass transfer co-efficient between two fluids given an added LFVP application has been experimentally verified by Hancil et al.. Further, Oberti et al., have shown enhanced mixing of particulate laden fluids in articulating channels (analogous to articulating blood vessels) secondary to external LFVP[39, 40]. Accordingly, LFVP mixing devices such as produced by Resodyn™ Acoustic Mixers (operable at a nominal frequency of 60 Hz) have found common use in industrial mixing of both fluids and solids.
A fourth notable finding in our experiment was that clot mobilization within the catheter segment was quite rare, and when occurring was also only observed in an LFVP group. While the mechanism for this is speculative, it is likely that the transmitted vibration simply caused disadhearment (or detachment) of the clot from along the catheter walls and the distal cap, which thereby allowed the clot to move freely within the agitated solution. There was no statistical difference in the likelihood of mobilization between SK enriched (1/8) and non-enriched (2/8) LFVP treatment groups.
A final notable finding to our experiment was that clot fragmentation (with division of clot into separated pieces) was also very rare, occurring only once in a SK non-enriched LFVP “HS only” group. Again while the mechanism for this is also speculative, it is likely that clot fragmentation may be simply a result of more extensive clot erosion with possibly an increased surface tension along the clot’s length during vibration induced lengthening and de-furlment. Inspection of LFVP treated clots (although more commonly in the SK enriched samples) did show aspects of extreme clot thinning into tiny strands – which amazingly were able to keep the clot intact. It is worth mentioning how strong these tiny strands (which are often as thin as a strand of spider’s web!) can be, being able to support the majority weight of the remainder of a clot when being held upright by the hand of an investigator (refer to Figure10).
Our findings raise the interesting question whether chest wall delivered LFVP in the low sonic frequency ranges (as opposed to, or maybe even working co-operatively with therapeutic ultrasound?) may hold potential as a knock out “one–two punch” in aid of clot disruptive, and more particularly IV thrombolytic therapy in treatment of STEMI. First by mechanically eroding clot surface layer and altering clot morphology to promote intra-luminal fluid channel development (which would enhance early flow and uncover fibrin binding sites), and second by facilitating diffusive bulk flow of clot disruptive drug agents from a concentration rich systemic circulation towards a concentration poor, otherwise stagnant, thrombosed coronary vessel.
It should be pointed out that transthoracic LFVP is prophesized to promote early clearance of acute coronary thrombosis by numerous additional mechanisms, which are beyond the scope of our present study.
First, it has been shown both clinically and in animal models that dLFVP when applied to the heart improves ischemic LV function, by improved myocardial relaxation, which has been attributed to enhanced diastolic filling leading to increased stroke volume by the Frank Starling mechanism[21, 28, 29]. This, by stabilizing a STEMI patient’s blood pressure (for example during heart failure or cardiogenic shock), would also augment systemic filtration pressures in assistance of bulk flow permeation of clot busters into a site of culprit thrombosis.
Second, improved myocardial relaxation by dLFVP would theoretically decrease coronary arteriolar and capillary flow resistance (by reduced diastolic myocardial compression) and is known to lower left ventricular (LV) diastolic pressures, which in combination should assist coronary flow and microcirculation during or following reperfusion.
Third, there is good evidence that LFVP carries potent vasodilatory capabilities for arteries in states of heightened vascular tone (i.e. by induced relaxation of the smooth muscles in the vessel wall)[41–43], and thus holds potential to induce direct coronary dilation at the site of acute coronary thrombosis, which often has (in at least 50% of STEMI cases) a degree of associated localized coronary spasm[44–47]. It is also worth mentioning that cyclic stress and strain exerted on an endothelial lining of an artery (which would likely be induced through a prophesized LFVP application)c, is predictive to cause liberation of beneficial mediators such as Nitric Oxide (NO) which is also a potent vasodilator. Indeed low frequency vibration stimuli has been experimentally shown to trigger NO release in various tissues[49–51]. LFVP’s vasodilatory mechanism in particular may therefore offer a degree of near immediate early re-flow in a high percentage of STEMI victims.
Fourth, LFVP should encourage disadhearement of acute thrombosis from the coronary wall (i.e. from site of ruptured plaque), and thereby promote very early recanlization of the major culprit epicardial vessel. External LFVP’s ability to clear clot from a stenosis site has been shown previously by our group, and has also been demonstrated by Folts et al. in reliable and immediate clearance of acute coronary and carotid thrombosis (albeit at a lower frequency, via direct hand tapping or shaking of the acutely thrombosed vessel) in open animal models.
Fifth, LFVP is known to stimulate endogenous liberation of fibrinolytic mediators[52, 53], thereby potentially offering a natural augmentation for localized fibrinolyis.
Six, LVFP should curtail stagnant arteriolar/capillary flow by prophylactic agitation of distal clotted fragments and by possibly lowering blood viscosity[54, 55], which may further assist in microcirculation during or following reperfusion. Researchers at Mt. Sinai Medical Centre (Miami, Florida) have recently studied the effects of low sonic vibration applied to the chest wall of rats which showed an up-regulation of endothelial derived NO Synthase (eNOS) with enhanced NO release, which in addition to its therapeutic vasodilatory properties has been discussed as a potential cardio protective mechanism in limitation of ischemic reperfusion injury.
There were several limitations to this present study. Firstly, the degree of removal of accessible fluid by our aspiration weighting technique could not be made identical between test runs. Great care however was taken to remove fluid using the same technique and in near identical fashion between samples, and the individual providing the post weighting was kept blinded to the form of treatment the clot was to, or had received. Secondly the meat slab was only a crude approximation of an overlying chest wall (lacking muscle fibers, ligaments, fat, fibrous pericardium, pericardial fluid and lung), and the underlying clotted vessel comprised a flexible catheter segment rather than a live coronary artery. However we felt that as an early experiment to assess LFVP’s ability to transmit and provide a basic clot disruptive effect over a thick chest wall sized attenuation barrier, this would at least provide a good “go–no go” hypothesis in that if LFVP produced a negligible effect the application would most certainly fail under less ideal clinical circumstances. It should be mentioned that LFVP has been shown to penetrate effectively from the human chest wall to the heart (even causing vibratory deformations to the deep posterior wall) by Koiwa and his associates by use of a more gentle vibration device (2 mm amplitude) than what we used in our experiment[21, 31]. Thirdly we had the advantage of localizing the vibrator placement upon the meat slab to ensure that the catheter segment underneath was vibrating, whereby this advantage would be absent clinically. However, the design of a vibratory attachment interface specific for inter-ribspace chest wall applications is presently underway which enables enough coverage over the base of the heart to maximize the chances that an LFVP source would generally overly a culprit coronary vessel. Fourthly, we cannot say by direct observation whether the relative improvement of remotely administered SK effectiveness with LFVP was due to a process of enhanced drug delivery (by facilitated diffusion), or simply an enhanced localized interaction of thrombolytic agent at the clot interface. The minimal degree of SK clot dissolution without LFVP however (virtually absent, only 3%) strongly suggests that the controls had very little if any contact with the SK molecule, so there very likely was at least a degree of enhanced drug delivery in the LFVP samples. Finally this study, because it was not a “flow model”, provided little information on whether LFVP may promote macro-embolization of clotted fragments downstream (or more worrisome upstream, especially in view of LFVP enhanced turbidity) which has been a concern to some colleagues. We can say however that while the remnant clots following LFVP treatment had always become thin and eroded in appearance, in only one rare occurrence (in 1 of 16 samples) was actual clot fragmentation observed (see Figure6). Hence the chances of a macro-embolic proximally situated piece of clot breaking off and flying out towards the systemic circulation would seem, at least from our study, unlikely. Clot fragments of course by common wisdom would generally tend to mobilize (if at all) downstream along with the vessels pressure gradient – to less harmful territory. In our experiment we did see LFVP promoted red color (representing dissolved, disaggregated, clot constituents) moving upstream towards and into the connecting line, and occasionally (in 3 of 16) samples there was a degree of upstream mobilization of the clot itself. However we point out that our system would not allow for anything but proximal clot mobilization (as distal flow was always blocked by the catheter segment’s distal cap), so no correlation can be drawn from our study whether there is an adverse tendency for LFVP to mobilize clot’s proximally versus more beneficially distally. Never-the-less LFVP induced proximal fluid turbidity warrants caution towards the potential of upstream liberation of coronary originated thrombo-emboli, hence any in- vivo study involving this prophesized application would require vigilant monitoring for stroke and acute peripheral arterial occlusion during safety analysis.
It is worth additional historical comment that our group has been placing faith in animal and human data from Koiwa and his associates (studies almost two decades old!) as gospel that dLFVP when applied to a human chest wall surface is sufficiently penetrative and causes enhanced Left Ventricular (LV) function and coronary flow in the ischemic heart. Koiwa’s work was abandoned in or about 1996, perhaps because of lack of funding, or because they were targeting use of their vibration device for treatment of heart failure (which probably would have been of little practical value).
The search for ideal LFVP parameters
It should be emphasized that our choice of studying “50 Hz” (versus any other low sonic or high infrasonic frequency) was based primarily on third party reports (Koiwa et al’s work) which attest to the therapeutic effect of diastolic timed 50 Hz on LV function and ability to enhance coronary flow[28–32]. Alternatively, we have found that 100 Hz, 0.5 mm LFVP offers clot clearing effects across a small 2 cm meat barrier, and Wobser et al. found that LFVP in the 50 500 Hz range disrupted big blood coagula in the stomach, with better efficacy at higher frequency levels. The ability for the heart to transmit received LFVP signals transversely (i.e. along arteries and the epi-myocardium) has been reported to generally occur at frequencies in the range of 20–120 Hz[23–26], so it is conceivable that any frequency within this range may comprise a reasonable choice in causing a therapeutic agitative effect to a coronary circulation. There is also the question about vibratory pattern. Engineers at Simon Fraser University have postulated that random or swept LFVP within a select frequency range may offer superior clot disruptive effects, such as to optimize turbulence, and ensure occasional striking of an optimal resonant clot disruptive excitation or cardiac frequency.
It is also possible that LFVP could be combined with therapeutic ultrasound with or without co-administration of ultrasonically active IV micro-bubbles (which oscillate in response to ultrasound, to augment acoustic agitation and intra-luminal sheer forces). Indeed this could be accomplished easily by simply mounting a therapeutic ultrasound transducer as a percussive contact (with or without an accompanying imaging system) upon an active end of a low frequency sonic actuator. In such an application it could be envisioned that LFVP would provide clot erosion and vasodilatory mechanisms to promote initial recanalization of a TIMI 0 flow vessel, which would thereby assist entry of systemic micro-bubbles (as well as thrombolytic drug agents), into a thrombosed culprit circulation. Much work in deciphering an optimal frequency, vibratory pattern, and potential use of multi frequency acoustic waveforms and adjunctive intravenous agents remains in development of this field.
The potential for external LFVP in the clearance of acute arterial thrombosis is far reaching beyond coronary syndromes. For example a more gentle LFVP application (i.e. to the cranium and/or neck of a patient) may be useful to expedite localized IV drug effectiveness in treatment of Acute Ischemic Stroke (AIS). In fact, Antic and his associates have demonstrated that headphone applied music (a gentle form of LFVP) enhances culprit Middle Cerebral Artery (MCA) blood flow in non thrombolysed AIS victims within a day following admission. External LFVP massage could also foresee ably be utilized in the acute emergency treatment of acute pulmonary emboli (particularly in saddle emboli – a life threatening condition) or acute peripheral arterial thrombosis as an alternative or bridge to emergency surgery or catheter based removal techniques. Clinical trials would be required on all these fronts.