Adipose derived stem cells and platelet rich plasma improve the tissue integration and angiogenesis of biodegradable scaffolds for soft tissue regeneration.

Current surgical reconstruction for soft tissue replacement involves lipotransfer to restore soft tissue replacements but is limited by survival and longevity of the fat tissue. Alternative approaches to overcome these limitations include using biodegradable scaffolds with stem cells with growth factors to generate soft tissue. Adipose derived stem cells (ADSCs) offer great potential to differentiate into adipose, and can be delivered using biodegradable scaffolds. However, the optimal scaffold to maximise this approach is unknown. This study investigates the biocompatibility of nanocomposite scaffolds (POSS-PCL) to deliver ADSCs with and without the addition of growth factors using platelet rich plasma (PRP) in vivo. Rat ADSCs were isolated and then seeded on biodegradable scaffolds (POSS-PCL). In addition, donor rats were used to isolate PRP to modify the scaffolds. The implants were then subcutaneously implanted for 3-months to assess the effect of PRP and ADSC on POSS-PCL scaffolds biocompatibility. Histology after explanation was examined to assess tissue integration (H&E) and collagen production (Massons Trichome). Immunohistochemistry was used to assess angiogenesis (CD3, α-SMA), immune response (CD45, CD68) and adipose formation (PPAR-γ). At 3-months PRP-ADSC-POSS-PCL scaffolds demonstrated significantly increased tissue integration and angiogenesis compared to PRP, ADSC and unmodified scaffolds (p < 0.05). In addition, PRP-ADSC-POSS-PCL scaffolds showed similar levels of CD45 and CD68 staining compared to unmodified scaffolds. Furthermore, there was increased PPAR-γ staining demonstrated at 3-months with PRP-ADSC-POSS-PCL scaffolds (p < 0.05). POSS-PCL nanocomposite scaffolds provide an effective delivery system for ADSCs. PRP and ADSC work synergistically to enhance the biocompatibility of POSS-PCL scaffolds and provide a platform technology for soft tissue regeneration.

Biomechanical characterisation of the human nasal cartilages; implications for tissue engineering.

Nasal reconstruction is currently performed using autologous grafts provides but is limited by donor site morbidity, tissue availability and potentially graft failure. Additionally, current alternative alloplastic materials are limited by their high extrusion and infection rates. Matching mechanical properties of synthetic materials to the native tissue they are replacing has shown to be important in the biocompatibility of implants. To date the mechanical properties of the human nasal cartilages has not been studied in depth to be able to create tissue-engineered replacements with similar mechanical properties to native tissue. The young's modulus was characterized in compression on fresh-frozen human cadaveric septal, alar, and lateral cartilage. Due to the functional differences experienced by the various aspects of the septal cartilage, 16 regions were evaluated with an average elastic modulus of 2.72 ± 0.63 MPa. Furthermore, the posterior septum was found to be significantly stiffer than the anterior septum (p < 0.01). The medial and lateral alar cartilages were tested at four points with an elastic modulus ranging from 2.09 ± 0.81 MPa, with no significant difference between the cartilages (p < 0.78). The lateral cartilage was tested once in all cadavers with an average elastic modulus of 0.98 ± 0.29 MPa. In conclusion, this study provides new information on the compressive mechanical properties of the human nasal cartilage, allowing surgeons to have a better understanding of the difference between the mechanical properties of the individual nasal cartilages. This study has provided a reference, by which tissue-engineered should be developed for effective cartilage replacements for nasal reconstruction.

Development of resorbable nanocomposite tracheal and bronchial scaffolds for paediatric applications.

BACKGROUND: Congenital tracheal defects and prolonged intubation following premature birth have resulted in an unmet clinical need for tracheal replacement. Advances in stem cell technology, tissue engineering and material sciences have inspired the development of a resorbable, nanocomposite tracheal and bronchial scaffold. METHODS: A bifurcated scaffold was designed and constructed using a novel, resorbable nanocomposite polymer, polyhedral oligomeric silsesquioxane poly(ϵ-caprolactone) urea urethane (POSS-PCL). Material characterization studies included tensile strength, suture retention and surface characteristics. Bone marrow-derived mesenchymal stem cells (bmMSCs) and human tracheobronchial epithelial cells (HBECs) were cultured on POSS-PCL for up to 14 days, and metabolic activity and cell morphology were assessed. Quantum dots conjugated to RGD (l-arginine, glycine and l-aspartic acid) tripeptides and anticollagen type I antibody were then employed to observe cell migration throughout the scaffold. RESULTS: POSS-PCL exhibited good mechanical properties, and the relationship between the solid elastomer and foam elastomer of POSS-PCL was comparable to that between the cartilaginous U-shaped rings and interconnective cartilage of the native human trachea. Good suture retention was also achieved. Cell attachment and a significant, steady increase in proliferation were observed for both cell types (bmMSCs, P = 0·001; HBECs, P = 0·003). Quantum dot imaging illustrated adequate cell penetration throughout the scaffold, which was confirmed by scanning electron microscopy. CONCLUSION: This mechanically viable scaffold successfully supports bmMSC and HBEC attachment and proliferation, demonstrating its potential as a tissue-engineered solution to tracheal replacement.

Nanotopography and plasma treatment: redesigning the surface for vascular graft endothelialisation.

INTRODUCTION: Vascular graft materials in clinical use, such as polytetrafluoroethylene (PTFE) and Dacron, do not endothelialise and have low patency rates. The importance of an endothelial cell layer on the luminal surface of a vascular graft is well-known with surface topography and chemistry playing an important role. The aim of this study was to investigate the potential of plasma treatment and topographical structures on the luminal graft surface to enhance the self-endothelialisation potential of a nanocomposite vascular graft. METHODS: POSS-PCU is a polycarbonate urea urethane (PCU) with a nanoparticle, polyhedral oligomeric silsesquioxane (POSS) incorporated within it. Planar, microgrooved, and nanopit patterned polymer films were fabricated using photolithography, electron beam lithography, reactive ion etching, and replication by solvent casting. Films were then exposed to oxygen plasma treatment at different powers for a fixed time (40 W, 60 W, 80 W/60 seconds). Effects of plasma treatment were assessed using scanning electron microscopy, atomic force microscopy and water contact angle analysis. Human umbilical vein endothelial cell (HUVEC) proliferation and morphology were characterised using immunostaining, live/dead staining, and Coomassie blue staining. RESULTS: Successful embossing of the micro- and nanostructures was confirmed. Oxygen plasma treatment of the different samples showed that increasing power significantly increased the hydrophilicity of the samples (p < .0001). Improved HUVEC adhesion was seen on plasma modified compared with untreated samples (p < .0001). Coomassie blue staining showed that after 5 days, cells started to form monolayers and live/dead staining showed the cells were viable. Immunostaining showed that HUVECs expressed nitric oxide synthase on all topographies with focal adhesions appearing more pronounced on nanopit surfaces, showing retention of morphology and function. CONCLUSION: These encouraging results indicate a future important role for plasma treatment and nanotopography in the development of endothelialised vascular grafts.

Luminal surface engineering, 'micro and nanopatterning': potential for self endothelialising vascular grafts?

OBJECTIVE: New technologies are being explored to meet the clinical need for an 'off-the-shelf' small diameter vascular graft with superior or at least equivalent properties to autologous vessel. The field of nanotechnology and fabrication promises major advances in biomaterial design and wall structure to deliver biomimetic grafts. This review brings together recent work on this topic. METHODS: A literature search was conducted of PubMed and ISI Web of Knowledge using relevant keywords. Articles published after January 2005 were given preference. Personal communications and PhD theses were also used as sources. RESULTS: An evolving focus on surface patterning of biomaterials has been found to carry great potential. Influencing cellular behaviour on prosthetic grafts using graft luminal surface modulation at the micro- and nano-levels is the basis of this recent concept in vascular graft development. CONCLUSION: This technology may deliver small diameter grafts with the potential for spontaneous in situ endothelialisation without the need for prior 'seeding', with the potential to open a new chapter in vascular graft development.

Surface and mechanical analysis of explanted Poly Implant Prosthèse silicone breast implants.

BACKGROUND: The recent events surrounding Poly Implant Prosthèse (PIP) breast implants have renewed the debate about the safety profile of silicone implants. The intentional use of industrial-grade instead of certified medical-grade silicone is thought to be responsible for reportedly higher frequencies of implant rupture in vivo. The differences in mechanical and viscoelastic properties between PIP and medical-grade silicone implant shells were investigated. Surface characterization of shells and gels was carried out to determine structural changes occurring after implantation. METHODS: Breast implants were obtained from women at the Royal Free Hospital (London, UK). PIP implants were compared with medical-grade control silicone implants. Tensile strength, tear resistance and elongation at break were assessed using a tensile tester. Surfaces were analysed using attenuated total reflectance-Fourier transform infrared (ATR-FTIR) spectroscopy. Spearman correlation analyses and Kruskal-Wallis one-way statistical tests were performed for mechanical data. RESULTS: There were 18 PIP and four medical-grade silicone implants. PIP silicone shells had significantly weaker mechanical strength than control shells (P < 0·009). There were negative correlations between mechanical properties of PIP shells and implantation times, indicative of deterioration of PIP shells over time in vivo (r(s) = -0·75, P = 0·009 for tensile strength; r(s) = -0·76, P = 0·001 for maximal strain). Comparison of ATR-FTIR spectra of PIP and control silicones demonstrated changes in material characteristics during the period of implantation suggestive of time-dependent bond breakage and degradation of the material. CONCLUSION: This study demonstrated an increased weakness of PIP shells with time and therefore supports the argument for prophylactic removal of PIP breast implants.

Tissue engineering of a hybrid bypass graft for coronary and lower limb bypass surgery.

Tissue-engineered blood vessels have largely relied on inelastic scaffolds or biological solutions with uncertain long-term in vivo durability. In this report we present for the first time a hybrid tissue-engineered bypass graft consisting of an elastic scaffold of compliant poly(carbonate-urea)urethane (CPU), incorporated with human smooth muscle cells (SMCs) and endothelial cells (ECs) from the same human source. Human vascular SMCs and ECs were extracted from umbilical cord vessels. The effect of shear stress preconditioning on cell retention on the hybrid bypass graft was investigated under pulsatile arterial flow conditions. Retention of ECs seeded onto CPU precoated with SMCs was significantly improved by a period of shear stress preconditioning, especially when the stress incrementally increased. This is probably because the mechanical stimuli orient cells and increase the release of matrix proteins and attachment factors. The stage is now set for developing a hybrid graft for in vivo studies.

Electrohydrodynamic jetting behaviour of polyhedral oligomeric silsesquioxane nanocomposite.

In this paper, we investigate in detail the electrohydrodynamic spraying of a nonbiodegradable nanocomposite polyhedral oligomeric silsesquioxane polymer developed in our laboratories and currently being explored for coating metallic stent materials. Different concentrations of the polymer have been dissolved to prepare, characterise, and electrohydrodynamically deposit the polymer on stainless steel. From the experiments, the solution containing 15 wt% polymer was selected for further investigation. The variation of film/coating thickness as a function of spraying time was studied and the structural features of the film were assessed using microscopy. Films were also tensile tested. This study has identified a process and conditions which can be used in our stent coating research.

Argon plasma surface modification promotes the therapeutic angiogenesis and tissue formation of tissue-engineered scaffolds in vivo by adipose-derived stem cells.

BACKGROUND: Synthetic implants are being used to restore injured or damaged tissues following cancer resection and congenital diseases. However, the survival of large tissue implant replacements depends on their ability to support angiogenesis that if limited, causes extrusion and infection of the implant. This study assessed the beneficial effect of platelet-rich plasma (PRP) and adipose-derived stem cells (ADSCs) on synthetic biomaterials in combination with argon plasma surface modification to enhance vascularisation of tissue-engineered constructs. METHODS: Non-biodegradable polyurethane scaffolds were manufactured and modified with plasma surface modification using argon gas (PM). Donor rats were then used to extract ADSCs and PRP to modify the scaffolds further. Scaffolds with and without PM were modified with and without ADSCs and PRP and subcutaneously implanted in the dorsum of rats for 3 months. After 12 weeks, the scaffolds were excised and the degree of tissue integration using H&E staining and Masson's trichrome staining, angiogenesis by CD31 and immune response by CD45 and CD68 immunohistochemistry staining was examined. RESULTS: H&E and Masson's trichrome staining showed PM+PRP+ADSC and PM+ADSC scaffolds had the greatest tissue integration, but there was no significant difference between the two scaffolds (p < 0.05). The greatest vessel formation after 3 months was shown with PM+PRP+ADSC and PM+ADSC scaffolds using CD31 staining compared to all other scaffolds (p < 0.05). The CD45 and CD68 staining was similar between all scaffolds after 3 months showing the ADSCs or PRP had no effect on the immune response of the scaffolds. CONCLUSIONS: Argon plasma surface modification enhanced the effect of adipose-derived stem cells effect on angiogenesis and tissue integration of polyurethane scaffolds. The combination of ADSCs and argon plasma modification may improve the survival of large tissue implants for regenerative applications.

Topical haemostatic agents.

BACKGROUND: A variety of local haemostatic agents is now available to stop troublesome bleeding. These agents are indicated for use during surgical interventions where conventional methods of haemostasis are not applicable because of the site of surgery or the degree of bleeding. METHOD: A literature search using the PubMed and ISI Web of Knowledge databases identified relevant studies on topical haemostatic agents. Manufacturers' recommendations were also sought through commercial websites. RESULTS AND CONCLUSION: A significant body of evidence now exists to support the use of topical haemostatic agents in a wide variety of clinical situations. The advantages and disadvantages of many of these agents are highlighted.

Increased risk of vascular events in systemic lupus erythematosus: is arterial stiffness a predictor of vascular risk?

Patients with systemic lupus erythematosus (SLE) have an increased vascular morbidity and mortality. Several established vascular risk factors are more prevalent in this population but cannot fully explain the reported excess atherosclerotic burden. Emerging vascular risk factors may also contribute to the increased vascular risk in these patients although the evidence is limited and often conflicting. SLE-specific risk factors also play a role in the pathogenesis of atherosclerosis.Given the multifactorial aetiology of vascular disease in SLE, an integrated index of risk could be useful in the management of these patients. Arterial stiffness possibly represents such an index and accumulating data suggest an increased prevalence of arterial stiffness in SLE. Many factors play a role in the loss of arterial elasticity in this population, including both emerging and established vascular risk factors. Arterial stiffness may emerge as a useful index for risk stratification in SLE and has the potential to guide therapeutic decisions in these patients.

Regional differences in the expression of nitric oxide synthase and specific receptors in the vascular tissues of control and diabetic rabbits: a pilot study.

BACKGROUND: Atherosclerosis can influence the expression of endothelial nitric oxide synthase (eNOS) as well as endothelin-1 (ET-1) and 5-hydroxytryptamine (5HT; serotonin) receptors. Diabetes has an effect on the onset, severity and pattern of atherosclerosis with a predilection for more distal arteries. We aimed to identify regional differences in the distribution of eNOS activity, ET-1 and 5HT receptors in vascular tissues obtained from control and diabetic rabbits. MATERIALS AND METHODS: The mid abdominal aorta, right renal and right femoral arteries were harvested from 12 adult rabbits (6 months old, 3-3.9 kg); 8 controls and 4 diabetic (induced using alloxan 7 months previously). Samples were stored in liquid nitrogen for Western immunoblotting for eNOS as well as ET-1 and 5HT receptors. RESULTS: Significant differences were found in the distribution of eNOS, ET-1 and 5HT between the aorta, renal and femoral arteries in the controls. The number of ET-1 receptors was significantly higher (aorta; p=0.016, renal; p=0.004, femoral; p=0.05,) whereas, the expression of eNOS was significantly lower (aorta; p =0.004, renal; p =0.004, femoral; p =0.008) when comparing arteries from normal rabbits with these from diabetics ones. The number of 5HT receptors was higher in arteries from diabetic rabbits but this was not statistically significantly. CONCLUSION: The "regional" distribution of eNOS activity as well as ET-1 and 5HT receptors in control rabbits varies significantly according to the vessel assessed. Further studies are needed to evaluate the effect of blocking these receptors (e.g. on the risk of re-stenosis). Regional receptor differences may explain why diabetes is linked with a predilection for atherosclerosis (and possibly calcification) in distal arteries.

Chemical group-dependent plasma polymerisation preferentially directs adipose stem cell differentiation towards osteogenic or chondrogenic lineages.

Human adipose derived stem cells (ADSCs) are being explored for the repair of craniofacial defects due to their multi-differentiation potential and ease of isolation and expansion. Crucial to using ADSCs for craniofacial repair is the availability of materials with appropriate biomechanical properties that can support their differentiation into bone and cartilage. We tested the hypothesis that different modifications of chemical groups on the surface of a nanocomposite polymer could increase human ADSC adhesion and selectively enhance their osteogenic and chondrogenic differentiation. We show that the COOH modification significantly promoted initial cell adhesion and proliferation over 14days compared to NH2 surfaces. Expression of focal adhesion kinase and vinculin was enhanced after plasma surface polymerisation at 24h. The COOH modification significantly enhanced chondrogenic differentiation as indicated by up-regulation of aggrecan and collagen II transcripts. In contrast, NH2 group functionalised scaffolds promoted osteogenic differentiation with significantly enhanced expression of collagen I, alkaline phosphatase and osteocalcin both at the gene and protein level. Finally, chorioallantoic membrane grafting demonstrated that both NH2 and COOH functionalised scaffolds seeded with ADSCs were biocompatible and supported vessel ingrowth apparently to a greater degree than unmodified scaffolds. In summary, our study shows the ability to direct ADSC chondrogenic and osteogenic differentiation by deposition of different chemical groups through plasma surface polymerisation. Hence this approach could be used to selectively enhance bone or cartilage formation before implantation in vivo to repair skeletal defects. STATEMENT OF SIGNIFICANCE: Human adipose derived stem cells (hADSCs) are an exciting stem cell source for regenerative medicine due to their plentiful supply and ease of isolation. However, the optimal environmental cues to direct stem cells towards certain lineages change have to has not been identified. We have shown that by modifying the surface of the scaffold with specific chemical groups using plasma surface polymerisation techniques we can control ADSCs differentiation. This study shows that ADSCs can be differentiated towards osteogenic and chondrogenic lineages on amine (NH2) and carboxyl (COOH) modified scaffolds respectively. Plasma polymerisation can be easily applied to other biomaterial surfaces to direct stem cell differentiation for the regeneration of bone and cartilage.

Biomechanical Characterisation of the Human Auricular Cartilages; Implications for Tissue Engineering.

Currently, autologous cartilage provides the gold standard for auricular reconstruction. However, synthetic biomaterials offer a number of advantages for ear reconstruction including decreased donor site morbidity and earlier surgery. Critical to implant success is the material's mechanical properties as this affects biocompatibility and extrusion. The aim of this study was to determine the biomechanical properties of human auricular cartilage. Auricular cartilage from fifteen cadavers was indented with displacement of 1 mm/s and load of 300 g to obtain a Young's modulus in compression. Histological analysis of the auricle was conducted according to glycoprotein, collagen, and elastin content. The compression modulus was calculated for each part of the auricle with the tragus at 1.67 ± 0.61 MPa, antitragus 1.79 ± 0.56 MPa, concha 2.08 ± 0.70 MPa, antihelix 1.71 ± 0.63 MPa, and helix 1.41 ± 0.67 MPa. The concha showed to have a significantly greater Young's Elastic Modulus than the helix in compression (p < 0.05). The histological analysis demonstrated that the auricle has a homogenous structure in terms of chondrocyte morphology, extracellular matrix and elastin content. This study provides new information on the compressive mechanical properties and histological analysis of the human auricular cartilage, allowing surgeons to have a better understanding of suitable replacements. This study has provided a reference, by which cartilage replacements should be developed for auricular reconstruction.

Endothelial progenitor cells and their potential clinical applications in peripheral arterial disease.

Endothelial progenitor cells (EPCs) were originally thought to be present only during embryonic development. New evidence suggests that they can persist into adult life, circulate in the peripheral blood and may play an important part in endothelial repair and replacement of dysfunctional endothelium. They may also play a role in the formation of new blood vessels (angiogenesis, vasculogenesis, and arteriogenesis) in ischemic tissues. In addition, EPCs have the potential to endothelialize small-diameter prosthetic vascular bypass grafts and generate a nonthrombogenic surface, thereby increasing the patency rate of these grafts. EPCs may also be used in the clinical assessment of risk of vascular disease. In this review, the authors discuss the potential use of EPCs in the management of peripheral arterial disease (PAD).

Effect of graded hypoxia on the rat hepatic tissue oxygenation and energy metabolism monitored by near-infrared and 31P nuclear magnetic resonance spectroscopy.

Alteration in hepatic cellular adenosine triphosphate (ATP) levels has been shown to be a sensitive index for hypoxic damage. Hepatic ATP metabolism can be monitored by 31P nuclear magnetic resonance (NMR). Near-infrared spectroscopy (NIRS) can measure tissue oxyhemoglobin (HbO2), deoxyhemoglobin (Hb), and cytochrome oxidase (Cyt Ox), which reflect ATP production. In this study, hepatic oxygenation parameters have been correlated with ATP metabolism under graded hypoxia. Sprague-Dawley rats underwent laparotomy for liver exposure. NIRS probes and an NMR coil were placed on the liver and the animal was positioned in the NMR magnet. Graded hypoxia was achieved by a stepwise reduction of the fraction of inspired oxygen (FiO2) from 15 to 4%. Recovery between the hypoxic periods was achieved using 30% oxygen. Hepatic tissue oxygenation parameters were measured continuously by NIRS; 31P-NMR spectra obtained at 1 min intervals from energy metabolites and intracellular pH were calculated. All the hypoxic grades produced an immediate reduction in HbO2 with a simultaneous increase in Hb. Cyt Ox was reduced significantly only with FiO2 of

Role of nanotechnology in development of artificial organs.

Improvements in our understanding of the interactions between implants and cells have directed attention towards nanoscale technologies. To date, nanotechnology has played a helping hand in the development of synthetic artificial organs and regenerative medicine. This includes the production of smart nanocomposite materials; fluorescent nanoparticles like Quantum Dots (QD) and magnetic nano particles (MNP) for stem cell tracking; and carbon nanotubes (CNT) and graphene for enhancement of material properties. The scope of this paper includes the role of nanoparticles in the development of nanomaterials; the chemical surface modifications possible to improve implant function and an overview of the performance of nano-engineered organs thus far. This includes implants developed for aesthetic purposes like nasal and auricular scaffolds, plastic and reconstructive surgical constructs (i.e. dermal grafts), hollow organs for cardiothoracic applications; and last but not least, orthopedic implants. The five-year outlook for nano-enhanced artificial organs is also discussed, highlighting the key research and development areas, available funds and the hurdles we face in accomplishing progression from prototypes on the laboratory bench to off-the-shelf products for the consumer market. Ultimately, this review aims to delineate the advantages of incorporating nanotechnology, as an individual entity or as a part of a construct for the development of tissue engineering scaffolds and/or artificial organs, and unravel the mechanisms of tissue cell-biomaterial interactions at the nanoscale, allowing for better progress in the development and optimization of unique nanoscale surface features for a wide range of applications.

Engineering of bypass conduits to improve patency.

For patients with severe coronary artery and distal peripheral vascular disease not amenable to angioplasty and lacking sufficient autologous vessels there is a pressing need for improvements to current surgical bypass options. It has been decades since any real progress in bypass material has reached mainstream surgical practice. This review looks at possible remedies to this situation. Options considered are methods to reduce prosthetic graft thrombogenicity, including endothelial cell seeding and developments of new prosthetic materials. The promise of tissue-engineered blood vessels is examined with a specific look at how peptides can improve cell adhesion to scaffolds.

The effect of graded steatosis on flow in the hepatic parenchymal microcirculation.

BACKGROUND: Steatosis is a major cause of microcirculatory impairment and graft dysfunction after liver transplantation. The mechanism of this circulatory compromise is unclear. The aim of this study was to evaluate in vivo the effect of steatosis on parenchymal microcirculation and on total hepatic blood flow in an animal model. METHODS: Four groups of New Zealand White rabbits (n=24) were investigated. Group 1 were fed on normal diet (controls). In groups 2, 3, and 4 graded steatosis was induced by feeding on a high cholesterol diet (1.5%) for 4, 8, and 12 weeks, respectively. After laparotomy and exposure of the liver, total hepatic blood flow (THBF) and the hepatic parenchymal microcirculation (HPM) were measured. These parameters were correlated with the degree of histological fat infiltration classified as mild (<30%), moderate (30-60%), or severe (>60%). RESULTS: The 4-, 8-, and 12-week cholesterol diets resulted in mild, moderate, and severe steatosis, respectively. There was an inverse correlation between the degree of fat infiltration and both HPM (Spearman r=-0.967, P<0.0001) and THBF (r=-0.893, P<0.0001). THBF was 137+/-6 ml/min in controls, which reduced to 121+/-3, 99+/-5, and 63+/-5 ml/min in steatotic livers of groups 2, 3, and 4, respectively. HPM was 226+/-5 flux units in the controls and 197+/-7, 119+/-8, and 37+/-9 flux units in steatotic livers of groups 2, 3, and 4 respectively. Comparing with controls using analysis of covariance, the fall in HPM and THBF was found to be significant (P<0.002) in the moderate and severe groups, but not significant (P>0.050) in the mild group. Parenchymal perfusion was reduced to a greater extent than total liver blood flow in moderate and severe grades of steatosis. CONCLUSIONS: Fatty infiltration reduces hepatic blood flow and parenchymal microcirculation. The latter is more markedly reduced with severe steatosis. This may explain the development of microcirculatory impairment and graft failure after transplantation of fatty livers despite adequate liver blood flow.