Genomic DNAs in a human leukemia cell line unfold after cold shock, with formation of neutrophil extracellular trap-like structures.

Cells are generally stored at low temperature which slows their cellular metabolism. However, the stress induced by cold shock can lead to cell injury or death. Here, we found that exposing human leukemia HL-60 cells to cold shock followed by rewarming (CS/RW) increased the number of dead cells with remodeled genomic structures in which DNA fibers fully unfold and extrude into extracellular space, similar to neutrophil extracellular traps (NETs). The unfolded DNA was associated with NET marker proteins, such as neutrophil elastase and histone H3, and could trap significant numbers of Escherichia coli. We also found that reactive oxygen species-a requisite for NET generation-accumulated during CS/RW in HL-60 cells. This treatment of HL-60 cells to trigger global DNA structural alterations has not been reported before, and helps to elucidate the mechanisms of human cellular response to cold stress.

Deficiency of interleukin-1 receptor antagonist promotes spontaneous femoral artery aneurysm formation in mice.

Femoral artery aneurysms (FAAs) are very rare, and their natural history is not well understood. In this study, we sought to analyze the pathogenesis of inflammatory FAAs in interleukin-1 receptor antagonist-deficient (IL-1Ra(-/-)) B6 mice. Systolic arterial pressures and plasma lipid levels of IL-1Ra(-/-) mice and wild-type (WT) mice did not differ significantly. However, IL-1Ra(-/-) mice spontaneously developed fusiform FAAs. Real-time PCR of 9-month-old IL-1Ra(-/-) mice revealed significantly increased mRNA levels of IL-1ß (6.6-fold), tumor necrosis factor-α (TNF-α) (12.4-fold), and matrix metalloproteinase-9 (6.0-fold) compared with WT mice. Histological analysis revealed numerous inflammatory cells around the FAAs in IL-1Ra(-/-) mice, and elastin staining showed destruction of both the internal and external elastic lamina in IL-1Ra(-/-) mice. Afterward, macrophage function was studied. After lipopolysaccharide (1 µg/mL) stimulation, IL-1Ra-deficient macrophages produced much higher levels of TNF-α than those from WT mice. Finally, we performed bone marrow cell transplantation. FAAs with many inflammatory cells in the adventitia were detected in several WT mice that received bone marrow cells from IL-1Ra(-/-) mice (44%), but not from WT mice (0%). Our study is the first to demonstrate that IL-1Ra deficiency in inflammatory cells disrupts immune system homeostasis and induces inflammatory FAAs in IL-1Ra(-/-) B6 mice. We believe that these mice will provide much information about the natural history and management of FAAs.

A diagnostic system for articular cartilage using non-destructive pulsed laser irradiation.

BACKGROUND AND OBJECTIVES: Osteoarthritis involves dysfunction caused by cartilage degeneration, but objective evaluation methodologies based on the original function of the articular cartilage remain unavailable. Evaluations for osteoarthritis are mostly based simply on patient symptoms or the degree of joint space narrowing on X-ray images. Accurate measurement and quantitative evaluation of the mechanical characteristics of the cartilage is important, and the tissue properties of the original articular cartilage must be clarified to understand the pathological condition in detail and to correctly judge the efficacy of treatment. We have developed new methods to measure some essential properties of cartilage: a photoacoustic measurement method; and time-resolved fluorescence spectroscopy. MATERIALS AND METHODS: A nanosecond-pulsed laser, which is completely non-destructive, is focused onto the target cartilage and induces a photoacoustic wave that will propagate with attenuation and is affected by the viscoelasticity of the surrounding cartilage. We also investigated whether pulsed laser irradiation and the measurement of excited autofluorescence allow real-time, non-invasive evaluation of tissue characteristics. RESULTS: The decay time, during which the amplitude of the photoacoustic wave is reduced by a factor of 1/e, represents the key numerical value used to characterize and evaluate the viscoelasticity and rheological behavior of the cartilage. Our findings show that time-resolved laser-induced autofluorescence spectroscopy (TR-LIFS) is useful for evaluating tissue-engineered cartilage. CONCLUSIONS: Photoacoustic measurement and TR-LIFS, predicated on the interactions between optics and living organs, is a suitable methodology for diagnosis during arthroscopy, allowing quantitative and multidirectional evaluation of the original function of the cartilage based on a variety of parameters.

The influence of Ho:YAG laser irradiation on intervertebral disc cells.

BACKGROUND AND OBJECTIVE: Various types of laser have been reported for percutaneous laser disc decompression (PLDD). The aim of this study was to understand the effects on intervertebral disc cells following Ho:YAG laser irradiation, using a three-dimensional culture model, and consider appropriate irradiation conditions. STUDY DESIGN/MATERIALS AND METHODS: Intervertebral discs from the lumbar spine were obtained from 36 female Japanese white rabbits and processed to obtain isolated cells in three-dimensional cultures. Photoacoustic and photothermal effects were investigated by irradiating three-dimensional cultures with Ho:YAG laser at 27 or 54 J. Residual cell counts after irradiation were estimated based on DNA content according to fluorometric assay. Lactate dehydrogenase levels were also investigated as a marker of damage to cell plasma membranes. Finally, proteoglycan synthesis was measured by rapid filtration assay of (35) S incorporation, as an index of matrix synthesis. RESULTS: Residual cell count tended to be higher in the 27-J group. Plasma membrane damage was higher and remained high longer after irradiation in the 54-J group. Proteoglycan synthesis was higher in the 27-J group than in the 54-J group, with some conditions (e.g., 90 mJ/pulse condition) showing marked activation of proteoglycan synthesis maintained for a long time after irradiation. CONCLUSIONS: Three-dimensional culture models of intervertebral disc cells are useful for clarifying relationships between cell reactions and photoacoustic and photothermal effects after laser irradiation. Total energy is closely related to optimization of irradiation conditions, which may allow optimization of cytoprotection and promotion of matrix synthesis in clinical practice.

Infrared imaging of an A549 cultured cell by a vibrational sum-frequency generation detected infrared super-resolution microscope.

We performed infrared (IR) spectroscopic imaging of molecular species in cultured cell interiors of A549 cells using in-house developed vibrational sum-frequency generation detected IR super-resolution microscope. The spatial resolution of this IR microscope was approximately 1.1 microm, which exceeds the diffraction limit of IR light. Therefore, we clearly observed differences in the signal intensity at various IR wavelengths which appear to originate from the differing IR absorptions of specific vibrational modes, and reveal the distribution of molecular species in the single cell. These results were never imaged with the conventional IR microscope.

Targeted increase in cerebral blood flow by transcranial near-infrared laser irradiation.

BACKGROUND AND OBJECTIVE: Brain function is highly dependent on cerebral blood flow (CBF). The precise mechanisms by which blood flow is controlled by NIR laser irradiation on the central nervous system (CNS) have not been elucidated. In this study, we examined the effect of 808 nm laser diode irradiation on CBF in mice. STUDY DESIGN/MATERIALS AND METHODS: We examined the effect of NIR irradiation on CBF at three different power densities (0.8, 1.6 and 3.2 W/cm(2)) and directly measured nitric oxide (NO) in brain tissue during NIR laser irradiation using an amperometric NO-selective electrode. We also examined the contribution of NO and a neurotransmitter, glutamate, to the regulation of CBF by using a nitric oxide synthase (NOS) inhibitor, N(g)-nitro-L-arginine methyl ester hydrochloride (L-NAME), and an N-methyl-D-aspartate (NMDA) receptor blocker, MK-801, respectively. We examined the change in brain tissue temperature during NIR laser irradiation. We also investigated the protection effect of NIR laser irradiation on transient cerebral ischemia using transient bilateral common carotid artery occlusion (BCCAO) in mice. RESULTS: We showed that NIR laser irradiation (1.6 W/cm(2) for 15-45 minutes) increased local CBF by 30% compared to that in control mice. NIR laser irradiation also induced a significant increase in cerebral NO concentration. In mice that received L-NAME, NIR laser irradiation did not induce any increase in CBF. Mice administered MK-801 showed an immediate increase but did not show a delayed additional increase in local CBF. The increase in brain tissue temperature induced by laser irradiation was estimated to be as low as 0.8 degrees C at 1.6 W/cm(2), indicating that the heating effect is not a main mechanism of the CBF increase in this condition. Pretreatment with NIR laser irradiation improved residual CBF and reduced the numbers of apoptotic cells in the hippocampus. CONCLUSION: Our data suggest that NIR laser irradiation is a promising experimental and therapeutic tool in the field of cerebral circulation research.

Efficacy of peritoneal oxygenation using a novel artificial oxygen carrier (TRM-645) in a rat respiratory insufficiency model.

PURPOSE: Supplemental oxygenation is essentially important in critically ill patients with potentially reversible pulmonary insufficiency. An extracorporeal membrane oxygenator and percutaneous cardiopulmonary support have been used for these patients. However, these techniques are associated with so many complications that an additional new therapeutic modality is required. The purpose is to investigate if the peritoneal cavity can be used as "extrapulmonary respiration" that is analogous to peritoneal dialysis and utilizes the efficacy of liposome-encapsulated hemoglobin (artificial oxygen carrier; TRM-645). METHODS: Rats weighing an average of 300 g (n = 18) received an incision in the right chest to generate pneumothorax, which resulted in severe and lethal hypoxia. Oxygenated TRM-645 and human red blood cells (MAP group) were administered into the peritoneum in the experimental rats' pneumothorax model. No treatment except the right pneumothorax was administered to the sham group. RESULTS: Survival times from the pneumothorax were significantly longer in the TRM-645 and MAP groups than in the sham group (32.0 +/- 6.9 and 22.0 +/- 4.9 min vs 9.2 +/- 1.9 min, P < 0.01). In addition, an arterial blood gas analysis showed that the oxygenation in levels significantly improved. CONCLUSIONS: The abdomen (peritoneum) can potentially become an "artificial lung" that can be employed in critical care settings. TRM-645 provides an alternative to the use of washed human red blood cells.

Amelioration of airway stenosis in rabbit models by photodynamic therapy with talaporfin sodium (NPe6).

It is difficult to treat patients with acquired airway stenosis, and the quality of life of such patients is therefore lowered. We have suggested the application of photodynamic therapy (PDT) as a new treatment for airway stenosis and have determined the efficacy of PDT in animal disease models using a second-generation photosensitizer with reduced photosensitivity. An airway stenosis rabbit model induced by scraping of the tracheal mucosa was administered NPe6 (5 mg kg(-1)), and the stenotic lesion was irradiated with 670 nm light emitted from a cylindrical diffuser tip at 60 J cm(-2) under bronchoscopic monitoring. PDT using NPe6 improved airway stenosis (P = 0.043) and respiratory stridor. A significant prolongation of survival time was seen in the PDT-treated animals compared to that in the untreated animals (P = 0.025) and 44% of the treated animals achieved long-term survival (>60 days). In conclusion, PDT using NPe6 is effective for improvement in airway stenosis.

Liposome-encapsulated hemoglobin transfusion rescues rats undergoing progressive hemodilution from lethal organ hypoxia without scavenging nitric oxide.

OBJECTIVE: To investigate the efficacy of liposome-encapsulated hemoglobin (LHb) transfusion in rats undergoing lethal progressive hemodilution. SUMMARY BACKGROUND DATA: Unlike other acellular hemoglobin-based oxygen carriers, LHb has lipid bilayer membranes that are similar to mammalian red blood cells (RBCs), which prevent hemoglobin from having any direct contact with the blood components and the endothelium. Acellular hemoglobin has a high affinity for nitric oxide (NO), and because they are reported to behave as NO scavengers, acellular hemoglobin-based oxygen carriers could have pressor effects on the peripheral vessels. During a massive hemorrhage, acellular hemoglobin caused vasoconstriction could decrease peripheral perfusion, thereby leading to diminished oxygen delivery. METHODS: Rats were subjected to blood withdrawal (0.2 mL/min) with a simultaneous resuscitation using an isovolemic fluid transfusion that contained LHb, 5% albumin, or washed rat RBCs for 150 minutes (n = 15 in each group). RESULTS: All rats transfused with LHb or RBCs were rescued from lethal progressive hemodilution, whereas none of the albumin-transfused rats survived. LHb did not affect the plasma NO metabolite levels, suggesting it was not a potent NO scavenger. LHb also improved hemodilution-induced metabolic acidosis, and reduced exaggerated neuroendocrine responses and injuries to the heart, liver, and kidney. It suppressed expression of hypoxia-inducible factor-1alpha in the liver and kidney, suggesting improvement of hypoxia at molecular response levels. However, neither transfused LHb nor RBCs improved the acute lung injury that occurs after progressive hemodilution. CONCLUSION: LHb transfusion is effective in rescuing rats undergoing progressive hemodilution from lethal organ hypoxia without scavenging NO.

Photodynamic therapy for airway stenosis in rabbit models.

BACKGROUND: Acquired airway stenosis in childhood is resistant to conventional treatment. We examined whether endoscope-assisted photodynamic therapy (PDT) is effective for airway stenosis in animal models of which the pathophysiologic progressions are similar to those of clinical cases showing rapid deterioration. METHODS: Tracheal mucosa-scraped rabbits were administered IV porfimer sodium (Photofrin; Wyeth K.K., Tokyo, Japan) [2 mg/kg], and the tracheal lesions were irradiated with 630 nm of light emitted from a cylindrical diffuser tip via a transtracheal approach. RESULTS: Rabbits without PDT (untreated animals) showed dense granulation tissue in the scraped lesion, resulting in airway stenosis complicated with respiratory stridor. PDT ameliorated the degree of airway stenosis (p = 0.008) and reduced respiratory stridor; rabbits that received PDT showed patchy granulation tissue that was only 20 to 30% of the volume of that seen in the untreated animals. Survival time of rabbits that received PDT was significantly prolonged compared with that of untreated animals (p = 0.03). CONCLUSIONS: PDT was effective for airway stenosis in rabbit models. This suggests that PDT has the potential as a new therapeutic method for airway stenosis originating from granulation tissue.

Simultaneous measurement of changes in light absorption due to the reduction of cytochrome c oxidase and light scattering in rat brains during loss of tissue viability.

We performed the simultaneous measurement of intrinsic optical signals (IOSs) related to metabolic activity and cellular and subcellular morphological characteristics, i.e., light scattering for a rat global ischemic brain model made by rapidly removing blood by saline infusion. The signals were measured on the basis of multiwavelength diffuse reflectances in which 605 and 830 nm were used to detect the IOSs that are thought to be dominantly affected by redox changes of heme aa(3) and CuA in cytochrome c oxidase (CcO), respectively. For measuring the scattering signal, the wavelength that was found to be most insensitive to the absorption changes, e.g., approximately 620 nm, was used. The measurements suggested that an increase in the absorption due to reduction of heme aa(3) occurred soon after blood clearance, and this was followed by a large triphasic change in light scattering, during which time a decrease in the absorption due to reduction of CuA occurred. Through the triphasic scattering change, scattering signals increased by 5.2 +/- 1.5% (n = 5), and the increase in light scattering showed significant correlation with both the reflectance intensity changes at 605 and 830 nm. This suggests that morphological changes in cells correlate with reductions of heme aa(3) and CuA. Histological analysis of tissue after the triphasic scattering change showed no alteration in either the nuclei or the cytoskeleton, but electron microscopic observation revealed deformed, enlarged mitochondria and expanded dendrites. These findings suggest that the simultaneous measurement of absorption signals related to the redox changes in the CcO and the scattering signal is useful for monitoring tissue viability in the brain.

Accumulation of photofrin in lesions of airway stenosis rabbit models.

Airway stenosis in childhood is resistant to conventional treatments. Endoscope-assisted photodynamic therapy (PDT) is a potent candidate for the therapeutic modality owing to the easy approach to the tracheal lesion and low degree of invasiveness. The aim of the present study was to examine whether a photosensitizer preferentially accumulates in the lesion of airway stenosis in order to explore the possible applicability of PDT. The tracheal mucosa of rabbits was scraped off, and the rabbits were intravenously administered with Photofrin. The tissue concentration of Photofrin was quantitatively measured by fluorometric analysis. Granulation formation was seen in the mucosa-deprived lesion, causing airway stenosis. Photofrin concentration in the granulation tissue was four-fold higher than that in the intact trachea and 10-fold higher than that in the liver, spleen, skin and muscle. Photofrin preferentially accumulated in the lesion of airway stenosis. A preliminary experiment on PDT using transtracheal illumination showed an amelioration of airway stenosis, resulting in reduction in respiratory stridor.

Effects of growth factors on heparin-carrying polystyrene-coated atelocollagen scaffold for articular cartilage tissue engineering.

The specific aim of our investigation is to study the potential use of a collagen/heparin-carrying polystyrene (HCPS) composite extracellular matrix for articular cartilage tissue engineering. Here, we created a high-performance extracellular matrix (HpECM) scaffold to build an optimal extracellular environment using an HCPS we originally developed, and an atelocollagen honeycomb-shaped-scaffold (ACHMS-scaffold) with a membrane seal. This scaffold was coated with HCPS to enable aggregation of heparin-binding growth factors such as FGF-2 and TGF-beta1 within the scaffold. Three-dimensional culture of rabbit articular chondrocytes within the HpECM-scaffold and subsequent preparation of a tissue-engineered cartilage were investigated. The results showed remarkably higher cell proliferative activity within the HpECM-pretreated-FGF-2 scaffold and the sustenance of phenotype within the HpECM-pretreated-TGF-beta1 scaffold. It was thought that both FGF-2 and TGF-beta1 were stably immobilized in the HpEMC-scaffold since HCPS generated an extracellular environment similar to that of heparan sulfate proteoglycan within the scaffold. These results suggest that an ACHMS-scaffold immobilized with HCPS can be a HpECM for cartilage regeneration to retain the heparin-binding growth factors within the scaffolds.

The effect of chitosan hydrogel containing DMEM/F12 medium on full-thickness skin defects after deep dermal burn.

Burn wound excision is considered necessary to prepare skin for grafting, and the success of graft "take" is thought to be dependent on the vascular supply to the wound. We previously showed that photocrosslinkable chitosan hydrogel containing DMEM/F12 medium (medium-Az-CH-LA) is a biocompatible and biodegradable biomaterial that promotes re-epithelialization and neovascularization. The current study was designed to determine the effect of medium-Az-CH-LA on deep dermal burn. Sixteen male Wistar rats were randomly divided into two groups that were treated with medium-Az-CH-LA (n=5) or a collagen sponge (n=5). Under anesthesia, the dorsal fur was shaved and the skin was exposed to water at 95 degrees C for 10s. After 2h, damaged tissue was removed from the fascia and dressed with medium-Az-CH-LA or a collagen sponge. Specimens were obtained after 2, 4, 6, 8, 12, 16 and 32 days for histological analysis. There was no significant difference in the time required for wound closure between the two groups, but the thickness of the granulation tissue in the medium-Az-CH-LA-treated group was greater than that in the collagen sponge-treated group. Moreover, degradation and neovascularization occurred earlier in the group treated with medium-Az-CH-LA compared with the collagen sponge-treated group. These findings suggest that early degradative and angiogenic activities of medium-Az-CH-LA may be beneficial for granulation tissue formation in deep dermal burn wounds.

Synthesis of spirocyclic C-arylribosides via cyclotrimerization.

[reaction: see text] Spirocyclic C-arylribosides were synthesized from the known gamma-ribonolactone derivative. Lithium acetylide addition followed by glycosylation with 3-(trimethylsilyl)propargyl alcohol converted the ribonolactone to silylated diynes. After desilylation or iodination, subsequent ruthenium-catalyzed cycloaddition of resultant diynes with alkynes or chloroacetonitrile gave spirocyclic C-arylribosides.

Polyion complex micelles for photodynamic therapy: incorporation of dendritic photosensitizer excitable at long wavelength relevant to improved tissue-penetrating property.

A polymeric micelle (DPcZn/m) system, which is formed via an electrostatic interaction of anionic dendrimer phthalocyanine (DPcZn) and poly(ethylene glycol)-poly(l-lysine) block copolymers (PEG-b-PLL), was prepared for use as an effective photosensitizer for photodynamic therapy. DPcZn/m exhibited strong Q band absorption around 650 nm, a useful wavelength for high tissue penetration. Dynamic light scattering studies indicated that the DPcZn/m system has a relevant size of 50 nm for intravenous administration. Under light irradiation, either DPcZn or DPcZn/m exhibited efficient consumption of dissolved oxygen in a medium to generate reactive oxygen species and an irradiation-time-dependent increase in photocytotoxicity. The photodynamic efficacy of the DPcZn was drastically improved by the incorporation into the polymeric micelles, typically exhibiting more than two orders of magnitude higher photocytotoxicity compared with the free DPcZn at 60-min photoirradiation.

Intracellular kinetics of ATX-S10.Na(II) and its correlation with photochemical reaction dynamics during a pulsed photosensitization process: effect of pulse repetition rate.

Although photodynamic therapy with pulsed light excitation has interesting characteristics, its photosensitization mechanism has not been fully elucidated. In this study, we showed that the intracellular kinetics of ATX-S10.Na(II), a lysosomal sensitizer, was closely related to photochemical reaction dynamics during photodynamic treatment of A549 cells with nanosecond pulsed light. Fluorescence microscopy revealed that at high frequencies of 10 and 30 Hz the sensitizer initially localized mainly in lysosomes but that it started to be redistributed to the cytosol in certain ranges of radiant exposures. These ranges were found to coincide with a regime of fluorescence degradation with limited oxygen consumption. On the other hand, at 5 Hz, there was no such a discontinuous behavior in the sensitizer redistribution characteristics throughout the period of irradiation; this was consistent with the fact that no reaction switching was observed. Two possible reasons for the appearance of the regime with limited oxygen consumption are discussed: participation of an oxygen-independent reaction and change in the microenvironment for the sensitizer caused by lysosomal photodamage. The pulse frequency-dependent intracellular kinetics of the sensitizer also explains our previous results showing higher cytotoxicity at 5 Hz than at 10 and 30 Hz.

Controlled releases of FGF-2 and paclitaxel from chitosan hydrogels and their subsequent effects on wound repair, angiogenesis, and tumor growth.

A photocrosslinkable chitosan (Az-CH-LA) aqueous solution resulted in an insoluble hydrogel like a soft rubber within 30 sec of ultraviolet light (UV)-irradiation. The photocrosslinked chitosan hydrogel showed strong sealing strength and potential use as a new tissue adhesive in surgical application. Paclitaxel, which is an anti-tumor reagent and a vascularization-inhibitor, retained in the photocrosslinked chitosan hydrogel, and were gradually released from the photocrosslinked chitosan hydrogel in vivo upon the degradation of the hydrogel. The paclitaxel-incorporated photocrosslinked chitosan hydrogels effectively inhibited tumor growth and angiogenesis in mice. On the other hand, the fibroblast growth factor (FGF)-2 molecules also retained in both the photocrosslinked chitosan and an injectable chitosan/IO(4)-heparin hydrogels, and were gradually released from the hydrogels upon their in vivo biodegradations. The activity of FGF-2 in the hydrogels was stable for long time (more than 14 days). The controlled release of biologically active FGF-2 molecules from the hydrogels caused an induction of the angiogenesis and, possibly, collateral circulation occurred in the healing-impaired diabetic (db/db) mice and the ischemic limbs of rats. The purpose of this review is to describe the effectiveness of the chitosan hydrogels (photocrosslinkable chitosan hydrogel and chitosan/IO(4)-heparin hydrogel) as a local drug delivery carrier for FGF-2 and paclitaxel to control wound repair, tumor growth, and angiogenesis. It is thus proposed that the chitosan hydrogels may be a promising new local carrier for drugs such as FGF-2 and paclitaxel.

Medium (DMEM/F12)-containing chitosan hydrogel as adhesive and dressing in autologous skin grafts and accelerator in the healing process.

Autologous skin grafts are considered necessary for the treatment of extensive skin defects. However, skin graft by suturing is a time-consuming medical handling and rather stressful event for recipients. To that end, tissue adhesives have been suggested in skin grafts. Chitosan hydrogel is well known as a wound dressing and tissue adhesive material showing biocompatibility, anti-infective activity, and the ability to accelerate wound healing. In this report, we evaluated the application of the chitosan hydrogel as a tissue adhesive in skin grafts. Although chitosan hydrogel shortened the operation time and resulted in a high graft absorption rate in comparison with suturing, wound epithelization was rather retarded. On the other hand, chitosan hydrogel was found more biocompatible than the commonly used tissue adhesive octyl-2-cyanoacrylate. When the chitosan hydrogel was premixed with a serum-free tissue culture medium DMEM/F12, it was found to easily degrade and promote wound epithelization. Histological examination revealed that the medium (DMEM/F12)-containing chitosan hydrogel was associated with the accumulation of polymorphonuclear leukocytes and neovascularization. In addition, immunohistochemical staining showed that the vascular endothelial growth factor (VEGF) was localized in the chitosan hydrogel degraded matrices. And infiltration of leukocytes determined the degradation activity with the D-glucose in the medium (DMEM/F12) suggested to play a central role in chitosan hydrogel degradation. Therefore, the medium (DMEM/F12)-containing chitosan hydrogel may become commonly accepted as a beneficial wound dressing and tissue adhesive in extensive wound management and skin grafts.

Bone formation using human adipose tissue-derived stromal cells and a biodegradable scaffold.

Human adipose tissue, obtained by liposuction, was processed to obtain a fibroblast-like population of cells or adipose tissue-derived stromal cells (ATSCs). The ATSCs, as well as bone marrow-derived mesenchymal stem cells (BMSCs), have the capacity for renewal and the potential to differentiate into multiple lineages of mesenchymal tissues. These cells are capable of forming bone when implanted ectopically in an appropriate scaffold. The aim of this study was to evaluate a beta-tricalcium phosphate (beta-TCP) as a scaffold and to compare the potential of osteogenic differentiation of ATSCs with BMSCs. Both cell types were loaded into beta-TCP disk and cultured in an osteogenic induction medium. Optimal osteogenic differentiation in ATSCs in vitro, as determined by secretion of osteocalcin, scanning electron microscope, and histology, were obtained in the culturing with the beta-TCP disk. Furthermore, bone formation in vivo was examined by using the ATSC- or BMSC-loaded scaffolds in nude mice. The present results show that ATSCs have a similar ability to differentiate into osteoblasts and to synthesize bone in beta-TCP disk as have BMSCs.