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Biomedical subjects

H Winet

Publications and source records attributed to H Winet.

At least 19 recordsLinked to original sources
PubMed2003

A bone fluid flow hypothesis for muscle pump-driven capillary filtration: II. Proposed role for exercise in erodible scaffold implant incorporation.

A model is presented for enhancement of fluid flow through bone matrix and any porous tissue engineering scaffold implanted within it. The mechanism of enhancement is the skeletal muscle pump in compartments adjacent to the bone. Pressure waves from muscle pump contractions aided by increased blood pressure during exercise coupled with temporary occlusion of arteries leading to and veins from the bone, increase hydraulic pressure in cortical bone capillaries so as to amplify capillary filtration. It is proposed that capillary filtration increase is sufficiently convective to contribute to bone fluid flow and associated percolation through tissue engineered scaffold matrix implants. Importance of this contribution is its relative role in maintaining seeded cells in bioreactor scaffolds. Validation of the hypothesis starts at a minimum level of demonstrating that capillary filtration is convective. At a maximum level confirmation of the hypothesis requires demonstration that capillary filtration-based interstitial flow is sufficient to stimulate not only host bone cells (as proposed in part I of the hypothesis) but bioreactor-seeded cells as well. Preliminary data is presented supporting the prediction that skeletal muscle contraction generates convective capillary filtration.

Animals↗
PubMed2001

Effect of polymethylmethacrylate particles on mature bone in the optical bone chamber.

Reaction of mature bone and its vasculature to 3.33 +/- 0.19 microm polymethylmethacrylate (PMMA) particles at a concentration of 2.5 x 10(8)/cc was measured using optical bone chamber implant intravital microscopy. Twelve adult female New Zealand White rabbits were divided into six receiving Healon alone (controls) and six receiving Healon plus PMMA. The particles were introduced to the bone chamber compartment after removing its overlying optical element, which was immediately reinstalled. Reaction was monitored weekly over a 6-week period using video and photographic imaging. Bone was labeled before treatment with oxytetracycline and after treatment with alizarin complexone. Perfusing blood was labeled with fluorescein isothiocyanate dextran-70 kDa (FITC-D70). Parameters measured were net bone resorption, from black and white images, bone turnover, from color images, vascularity, and average vessel caliber. Neither bone turnover nor vessel caliber were significantly affected at the p < or = 0.050 level over time. In contrast, bone resorption was significantly greater and vascularity significantly less in the presence of PMMA. It was inferred that any differences in bone turnover were masked by resorption of new bone. It was concluded that the lack of a PMMA effect on average vessel caliber meant that the vascularity effects were not due to angiogenesis, but to vessel recruitment (or its opposite), an effect more consistent with inflammation than repair. The lack of vascularity increase in PMMA-treated compartments also suggested that increased resorption was a local phenomenon, because blood supply had not increased to provide the extra osteoclasts required for observed net bone loss.

Animals↗
PubMed2001

Evidence for reperfusion injury in cortical bone as a function of crush injury ischemia duration: a rabbit bone chamber study.

A model for critical limb ischemia was produced by occluding femoral vessels in 24 rabbits with a pneumatic cuff for 0, 2, 4, or 6 hours. Immediate sequelae and subsequent creeping substitution of cortical bone were observed in vivo using an implanted tibial window, the optical bone chamber implant (with intravital microscopy), and then by light and fluorescence microscopy of fluorochrome-labeled and surface-stained ground sections of retrieved implants. Six rabbits were used as controls (0 h) for each ischemia treatment, and the animals were monitored for 5 weeks postocclusion. A subpopulation of 13 implants was retrieved after euthanization and then histologically assessed for bone necrosis and remodeling. The hypothesis tested was that reperfusion injury during the 24 h after occluder release (reperfusion phase), and vessel perfusion/caliber, angiogenesis, and net bone resorption during the 5 subsequent weeks (creeping substitution phase), would exhibit ischemia duration-dependent effects. All animals could bear weight on the affected limb to ambulate by 1 week posttreatment. Two-way analysis of variance (ANOVA) comparison of the resulting data confirmed a significant difference between control and ischemia-treated rabbits for: (1) vessel perfusion/reperfusion; (2) vessel caliber; and (3) net bone resorption. Vascular responses to 4 vs. 6 h of ischemia were not significantly different, but net bone resorption was strictly ischemia duration-dependent. The conclusion that reperfusion injury was the mechanism spreading ischemia to more vessels was supported by a decrease in reperfusion and caliber of vessels, and an increase in vascular permeability and leukocyte adherence during the reperfusion phase. It is postulated that reperfusion injury produces a secondary ischemia that amplifies the occlusion-created primary ischemia and, in the present work, may have been succeeded by progressive episodes of ischemia, similar to the infarction pattern of ischemic hearts.

Animals↗
PubMed1998

Fibroblast growth factor-2 alters the effect of eroding polylactide-polyglycolide on osteogenesis in the bone chamber.

The effects of recombinant human fibroblast growth factor-2 (rhFGF-2) in the presence of eroding 50:50 poly(DL-lactide-co-glycolide) (PDLLG) on acute bone healing were studied in the optical bone chamber (BCI). BCIs were loaded with disks of PDLLG surrounded by one of four rhFGF-2 doses. Fifty-two female rabbit right tibias were implanted. Commencing the third week post implantation (W3) healing in the BCI compartment was observed weekly, using intravital microscopy, until W8. The doses were: unloaded, loaded with polymer only, and polymer plus 0.5, 1.0, and 10 microg rhFGF-2. Videotaped and photographed bone images were measured and analyzed using a frame-grabber digitizing system. Comparison with controls revealed that ossification rates were significantly above normal in rabbits loaded with polymer plus any of the rhFGF-2 doses. Comparison with polymer-only BCIs showed that PDLLG plus any of the three rhFGF-2 doses was linked with ossification rates significantly higher than baseline. The results indicated that FGF-2 in the dose range studied effectively can overcome the retarding effects of eroding polymer on ossification that has been reported by this laboratory. Interpretation of the retarding effects of the polymer disks, although consistent with previously studied washer-shaped devices of the same material, was complicated by a difference in erosion rate. This result supports the notion that erodible device geometry is a major factor in determining biocompatibility and must be considered in the design of carriers. Accordingly, programming of dose specificity for delivering a given polypeptide cytokine to a given host site must allow for the inhibitory effects of an eroding carrier and the influence of device geometry on these effects and erosion.

Animals↗
PubMed1998

Approaches to study of ischemia in bone.

Ischemic osteonecrosis is a complication of certain traumatic and a number of idiopathic conditions. The course of the disease may result in collapse of the convex member of a joint and osteoarthritis, often requiring arthroplasty. Increasing incidence in young adults poses a challenge for development of long-term joint prostheses. Current status of research into the disease is discussed and three new models using intravital microscopy described. The first, an arterial occlusion (AO) model, creates ischemia by occluding the common iliac artery exclusively, avoiding direct trauma on other tissues in the limb. The second, a total occlusion (TO) model utilizes classical tourniquet occlusion of the thigh vessels. The third, a venous occlusion (VO) model, is also a tourniquet procedure but it blocks occlusion of the femoral artery with a protective sheath. Preliminary results from AO and TO studies are reported which show that reperfusion injury is detectible after ischemia doses as short as 4 h. This complication was confirmed by observation of leukocyte adherence, secondary ischemia, and abnormal vessel leakage. Also, a new quantitation of osteonecrosis is introduced whereby fluorescently-tagged dead osteocytes and computer-based image processing provide values for an "osteonecrosis index." Viewing of all vascular events is made possible by intravital microscopy through a bone chamber window implanted in rabbit tibias. It is proposed that such chronic visual techniques allow quantitation of events leading to osteonecrosis as well as the revascularization, resorption and bone apposition of creeping substitution which characterizes postischemia recovery.

Animals↗
PubMed1997

Comparative bone healing near eroding polylactide-polyglycolide implants of differing crystallinity in rabbit tibial bone chambers.

Eroding poly(DL-lactide-co-glycolide) (PDLLG) washers and poly(L-lactide-co-glycolide) (PLLG) threads were observed chronically in vivo following loading in a bone chamber tibial implant (BCI). Images were recorded using intravital microscopy of the implanted rabbit. Erosion and bone healing, as represented by angiogenesis and osteogenesis, was determined from changes in projected area of observed polymer, vessels and bone, respectively. Erosion rates of the two polymers were significantly different. Healing adjacent to both polymers differed significantly from controls. Healing response to each polymer was also different, with the faster eroding PDLLG causing more deviation from normal osteogenesis and angiogenesis than did PLLG. It was speculated that the faster eroding polymer released macrophage-stimulating fragments earlier in the healing process, thus altering the normal macrophage-endothelial cell interaction which in turn affected angiogenesis-linked components of osteogenesis.

Analysis of Variance↗
PubMed1996

Acidity near eroding polylactide-polyglycolide in vitro and in vivo in rabbit tibial bone chambers.

Eroding poly(DL-lactide-co-glycolide)(PDLLG) washers were observed chronically in vitro and in vivo following loading in a bone chamber tibial implant (BCI). Images were recorded using intravital microscopy of the implanted rabbit and direct pH measurements were obtained of the tissue in the bone chamber using a combination probe-reference microelectrode. While statistically significant pH differences were found between the control (unloaded) and experimental (PDLLG-bearing) BCIs, they were only of the order of 0.2 pH units. This value proved to be physiologically insignificant as no statistically significant difference in bone defect healing, as indicated by angiogenesis, was detected. It was shown that the measured small pH changes during PDLLG washers erosion would result whether the buffer was phosphate-buffered saline replaced weekly or interstitial fluid subject to vascular exchanges.

Animals↗
PubMed1996

The role of microvasculature in normal and perturbed bone healing as revealed by intravital microscopy.

Bone chamber intravital microscopy brings to the study of bone circulation a combination of the control volume of in vitro models and the chemical complexity of in vivo models. As an optical tool it provides a window to circulatory events at the tissue level of magnification. In particular, it allows measures of microvascular physiology 1) in space by magnifying local perfused vasculature and microcirculation at any instant and 2) in time by providing the same volume of tissue for weekly viewing of an evolving process such as bone healing. This hollow screw's windows have revealed: 1) a consistent order for vascular and bone progression during healing, 2) vascular changes in response to implanted polymers and 3) preliminary data about effects of hyperbaric oxygenation and pulsed electromagnetic fields on vascular aspects of healing. The parameters measured are osteogenesis, angiogenesis, blood supply and permeability.

Animals↗
PubMed1995

Incorporation of polylactide-polyglycolide in a cortical defect: neoangiogenesis and blood supply in a bone chamber.

Erodible polymers are an alternative to metals for fracture fixation (for example, in the malleolus) and for maxillofacial reconstruction. In this study, the vascular response to eroding polylactide-polyglycolide copolymer threads was observed chronically in a bone chamber implant, with use of intravital microscopy. A bone chamber implant loaded with 100 microns thick polylactide-polyglycolide threads was implanted into the right tibia in 15 mature female New Zealand White rabbits. Periodic intravital microscopic observations were performed from the third to the tenth or twelfth week after implantation. Vascularization, blood flow, and trabecular growth into the chambers from the medial cortex were recorded on videotape and analyzed using digital image processing. A statistically significant delay of neo-osteogenesis in the presence of this copolymer was described in an earlier report. The present report describes the measures of neoangiogenesis and blood supply; there was a significant delay in neoangiogenesis. It is suggested that both delayed angiogenesis and osteogenesis were secondary consequences of the macrophage response to slowly eroding poly-L-lactide crystal nanoparticles and the influence of reduced nutrient exchange. The lesser effect on blood supply and vascular volume fraction was seen to be linked to the slowing down of angiogenesis, as the latter allowed vessels to mature, with a widening of their calibers. This homeostatic adjustment was interpreted as being only partially successful in restoring control levels of oxygen delivery, because resulting increases in vessel surface area did not reach control levels. Thus, in the presence of eroding polylactide-polyglycolide, the oxygen supply and extravasation of other nutrients may be below normal during healing phases when the need is critical.

Analysis of Variance↗
PubMed1993

Incorporation of polylactide-polyglycolide in a cortical defect: neoosteogenesis in a bone chamber.

One hundred-micron-thick threads of 40-kDa copolymer polylactide:polyglycolide (PLGA) in a 50:50 molar ratio in bone chambers were implanted in rabbit tibias. Weekly intravital microscopic observations were used to assess trabecular ingrowth and PLGA absorption, processes referred to as incorporation. Nineteen rabbits were observed from the 3rd (W3) to the 10th or 12th (W10/W12) week postimplantation. Analysis of digitally processed video images obtained weekly showed that PLGA absorption was completed in about 8-10 weeks and trabecular ingrowth delayed to about 1 week slower than its control value. Four of the bone chambers showed PLGA unraveled by week 3, no trabecular ingrowth, and significantly reduced neovascularization. No such apparent inhibitory effects were observed in bone chambers that displayed PLGA absorption. No causal link could be established between the rate of neoosteogenesis and the rate of PLGA absorption, but it was clear that incorporation slowed and rapid degradation inhibited bone regeneration. It was concluded that copolymer synthesis and postsynthesis processing must result in a synthetic with a predictable micro- and macro-molecular structure before one can predict its incorporability in a given reconstruction or fixation site.

Absorption↗
PubMed1990

A control model for tibial cortex neovascularization in the bone chamber.

Neovascularization across a gap defect in a rabbit tibial cortex was monitored using the optical bone chamber implant (BCI). Cortical bone growing by apposition as trabeculae was observed weekly as it penetrated a slit into a tissue space in vivo and in situ. Each rabbit was viewed weekly with an intravital microscope from 3 to 8 weeks postimplantation. The constant field of view was the slit-gap tissue space, which was 100 microns thick and 2 mm in diameter. Vessels were imaged with epi-illuminated fluorescence microscopy as they carried FITC-dextran 70 that had been injected into an aural vein. Observations were videotaped and photographed. Videotape frames were analyzed with a digital image processing system to obtain measures of vessel length per unit volume (L/V) of fibroblastic granular tissue and trabeculae, caliber C, and flow velocity u, all as functions of time. Observations supported the conclusions that (1) neovascularization precedes neo-osteogenesis, (2) major vessels tend to align with the tibial axis, (3) bone apposition-generated destruction of fibrous granular tissue vessels stimulates fibrous granular tissue angiogenesis, which keeps its L/V constant, (4) L/V in trabeculae increases with time, and (5) blood supply (Q) and nutrient exchange in healing trabeculae are not positively correlated. Thus, O2 supply to the trabeculum cannot be predicted from Q alone because the nutrient exchange area is not constant. It was noted that an increase in the potential nutrient exchange area occurred in both fibrous granular tissue and osseous vessels and the volume fraction of blood decreased in the fibrous granular tissue and remained constant in the trabeculae.

Animals↗
PubMed1990

Neo-osteogenesis of haversian trabeculae through a bone chamber implanted in a rabbit tibial cortex: a control model.

Neo-osteogenesis of cortical bone trabecular was observed as they regenerated into a bone chamber implant by appositional growth. Measures of change in bone area were obtained from 13 rabbits each week starting the third and ending the eighth week postimplantation. Observations were made using intravital microscopy and were analyzed using digital image processing. Images were computer-captured video frames equivalent to 78 (6 x 13) separate observations. They were measured by tracing the trabecular outlines with a digitizing crosshair each week and comparing changes in area as a percent of the circular field-of-view ("slit-gap") filled. Data supported the hypothesis that trabecular regeneration week 3 to week 8 followed the logistic growth curve regression: A% 100%/1 + 9.47e-0.7747(t-3) where A% is the percent of slit-gap area covered by bone and t is time, at a very high significance level. Nevertheless, a highly significant linear regression fit the data. Statistical analysis showed that the regression line could be fit to bone area measurements from weeks 3 to 8 (W3-W8) postimplantation, giving a constant neo-osteogenesis rate of 7.42 +/- 0.67 X 10(4) microns 2/day and a decreasing linear neo-osteogenesis rate from 73 microns/day at W4 to 21 microns/day at W8; the latter is based on a circle-segment approximation of trabeculum shape. This range approximated a bridge between ranges for cortical gap defect healing and porous ingrowth healing reported by other workers and supported the hypothesis that the BCI control model was a cross between gap healing and porous ingrowth.

Air Pressure↗
PubMed1989

A horizontal intravital microscope-plus-bone chamber system for observing bone microcirculation.

A horizontal intravital microscope has been built which utilizes the telescope optics format with infinity-corrected objectives. The intravital stage accomodates a rabbit into which a tibial bone chamber has been implanted. The chamber is itself an optical component and is considered an integral part of the system. Each animal is observed in supine position with its implanted leg held in an adjustable spring-loaded yoke which aligns the bone chamber in the optical axis. Long working-distance objectives and condensers allow for viewing of the slit-gap tissue in the bone chamber with both transmitted and epi-illumination; the latter being applied for fluorescence studies. A unique feature of the system is a transfer lens holder which can be relocated within the microscope tube. This adjustable lens holder allows for changing of transfer lenses, thereby freeing the investigator to utilize objective lenses with preferred numerical apertures for a given image size.

Animals↗
PubMed1988

Wound healing in the bone chamber 1. Neoosteogenesis during transition from the repair to the regenerative phase in the rabbit tibial cortex.

The optical bone chamber technique that includes intravital microscopy is described and is then applied to measuring primary wound healing neoostogenesis in rabbit tibia cortical bone during the period when fibrovascular tissue is being replaced by regenerating osteovascular tissue. The small population of rabbits sampled allowed only tentative conclusions. The quantitative measurements, a direct from of histomorphometry, are applied to determine the consistency of observations, with the hypothesis that healing into the bone chamber slit-gap follows the pattern of primary bone regeneration established as "characteristic" by other studies. The results supported the hypothesis and showed that bony ingrowth started during the third postoperative week (W3) with a maximum linear growth rate of 85.5 micron/day for a bone front. Evidence for remodeling by W6 was also obtained. While the bone chamber environment for tissue ingrowth is artificial, it can generate quantitative data that may provide a statistically valid basis for modeling pathophysiologic processes associated with bone wound healing.

Animals↗
PubMed1987

The role of the periciliary fluid in mucociliary flows: flow velocity profiles in frog palate mucus.

Observations of flow velocity profiles over frog mucociliated palate are used to estimate viscosity, shear rate and shear stress in the periciliary flow field. The ability of cilia to generate significant shear stress at long distances and their utility as rhoeometers are examined. It is proposed that the depth of significant ciliary shear penetration into the periciliary fluid is sufficient to move mucus masses well beyond the ciliary tips, obviating the need for tip penetration where anchoring phenomena are sufficiently reduced.

Animals↗
PubMed1984

On the mechanics of mucociliary flows. III. Flow-velocity profiles in frog palate mucus.

Flow-velocity profiles over excised frog ciliated epithelium were obtained for the region within about 600 micron of the mucosa. Fluorescent particles were used as flow tracers. Both a control and an autologous mucus suspension were observed. The control culture medium was bounded by the walls of the observation chamber, and mucus was deposited on the epithelium as a blob after mixing it with tracers. In spite of the difference in boundary conditions the two profiles, normalized to maximum particle velocity and solution depth, were indistinguishable at heights over 60 micron from the mucosa. The near-mucosa profiles in contrast were unalike with mucus exhibiting a greater shear gradient than the control culture medium. It was concluded that ciliary contact is not necessary for generation of mucus flow provided the ciliary shear is not negated by the mucus "flake" or "slab" being in simultaneous contact with significant ciliostatic patches which would act as anchors.

Animals↗
PubMed1984

Observations on the response of human spermatozoa to gravity, boundaries and fluid shear.

Human sperm motility response to three mechanical stimuli, gravity, fluid flow shear and rigid boundaries, was measured in a tube of 310 X 400 microns calibre. Data were gathered by cine recordings at various focussing levels d across the tube and analysed with a computerized image analysis system. The most influential stimulus was the tube wall near (more than 'at') which the swimmers tended to accumulate, leaving the fluid beyond 100 microns from the wall (d = 100) vacant of motile spermatozoa. The boundary effect was evident as soon as the spermatozoa could be viewed after loading, and accumulation, measured as frequency, as a function of d did not change with time t. This response was not significantly altered by the addition of laminar flow with a centre line velocity of about 400 microns/sec. In flow shear, spermatozoa aligned positively (in the flow direction) at the wall but negatively by about 30 microns from the wall where the velocity gradient (= shear rate) was about 3.5 sec-1. The response to gravity was relatively weak with 11 spermatozoa positive (swimming downwards) for each 9 negative. Neither the boundary effect nor the 'rheotaxic' effect were influenced by gravity as there was no statistical difference in orientation or distribution patterns between vertically and horizontally flowing suspensions. It is suggested that the boundary effect cannot be ignored in in-vitro manipulations, particularly when spermatozoa are observed or extracted. Its importance in vivo lies in the degree to which the tubes transporting motile spermatozoa seem to have mechanisms for reversing the wall accumulation tendency.

Computers↗