Search

Cellular stress, combined with dysfunctional, inadequate mitochondrial phosphorylation, produces an excessive amount of reactive oxygen species (ROS) and an increased level of ROS in cells, which leads to oxidation and subsequent cellular damage. Because of its cell damaging action, an association between anomalous ROS production and disease such as Type 1 (T1D) and Type 2 (T2D) diabetes, as well as their complications, has been well established. However, there is a lack of understanding about genome-driven responses to ROS-mediated cellular stress. Over the last decade, multiple studies have suggested a link between oxidative stress and microRNAs (miRNAs). The miRNAs are small non-coding RNAs that mostly suppress expression of the target gene by interaction with its 3’untranslated region (3′UTR). In this paper, we review the recent progress in the field, focusing on the association between miRNAs and oxidative stress during the progression of diabetes.

The chemical behavior of various oligoenes 2 has been studied. The catalytic hydrogenation of diene 3 yielded monoene 4. Triene 7 was hydrogenated to diene 8, monoene 9 and saturated hydrocarbon 10. Bromine addition to 3 and 7 yielded the dibromides 17 and 18, respectively, i.e., the oligoene system has been attacked at its terminal olefinic carbon atoms. Analogously, the higher vinylogs 19 and 20 yielded the 1,8- and 1,10-bromine adduts 23 and 24, respectively, when less than 1 equivalent of bromine was employed. Treatment of tetraene 19 with excess bromine provided tetrabromide 25. In epoxidation reactions, both with meta-chloroperbenzoic acid (MCPBA) and dimethyldioxirane (DMDO) two model oligoenes were studied: triene 7 and tetraene 19. Whereas 7 furnished the rearrangement product 31 with MCPBA, it yielded the symmetrical epoxide 32 with DMDO. Analogously, 19 was converted to mono-epoxide 33 with MCPBA and to 34 with DMDO. Diels–Alder addition of 7 with N-phenyltriazolinedione (PTAD) did not take place. Extension of the conjugated π-system to the next higher vinylog, 19, caused NPTD-addition to the symmetrical adduct 37 in good yield. Comparable results were observed on adding NPTD (equivalent amount) to pentaene 20 and hexaene 21. Using 36 in excess provided the 2:1-adduct 40 from 21 and led to a complex mixture of adducts from heptaene 22. With tetracyanoethylene (TCNE) as the dienophile, tetraolefin 19 yielded the symmetrical adduct 43, although the reaction temperature had to be increased. Pentaene 20 and hexaene 21 led to corresponding results, adducts 44 and 45 being produced in acceptable yields. With nonaene 42 and TCNE the 2:1-adduct 48 was generated according to its spectroscopic data. Exploratory photochemical studies were carried out with tetraene 19 as the model compound. On irradiation this reacted with oxygen to the stable endo-peroxide 52.

Substantial amounts of nonendocrine cells are implanted as part of human islet grafts, and a possible influence of nonendocrine cells on clinical islet transplantation outcome has been postulated. There are currently no product release criteria specific for nonendocrine cells due to lack of available methods. The aims of this study were to develop a method for the evaluation of pancreatic ductal cells (PDCs) for clinical islet transplantation and to characterize them regarding phenotype, viability, and function. We assessed 161 human islet preparations using laser scanning cytometry (LSC/iCys) for phenotypic analysis of nonendocrine cells and flow cytometry (FACS) for PDC viability. PDC and beta-cells obtained from different density fractions during the islet cell purification were compared in terms of viability. Furthermore, we examined PDC ability to produce proinflammatory cytokines/chemokines, vascular endothelial growth factor (VEGF) and tissue factor (TF) relevant to islet graft outcome. Phenotypic analysis by LSC/iCys indicated that single staining for CK19 or CA19-9 was not enough for identifying PDCs, and that double staining for amylase and CK19 or CA19-9 allowed for quantitative evaluation of acinar cells and PDC content in human islet preparation. PDC showed a significantly higher viability than beta-cells (PDC vs beta-cell: 75.5+/-13.9 and 62.7+/-18.7%; P

As shown in this study, the solvatochromic behavior of polyenes depends exclusively on the polarizability of the medium and, even more interestingly, their solvatochromism increases markedly with increasing length of the polyene chain. By virtue of the electronic nature of the interaction of polyenes with the medium, their solvatochromic response to a polarizability change is instantaneous, making these compounds extremely effective polarizability probes for molecular environments. The extreme sensitivity of polyenes to the polarizability of their environment is consistent with the fact that changes in molecular architecture such as those occurring in photosynthetic systems can give rise to polarizability gradients resulting in red shifts in the 1Ag --> 1Bu transition, thereby opening up new channels directing the energy transfer involved to energy trapping sites in such systems.

Hepatic homeostasis is essential for survival in critically ill and burned patients. Insulin administration improves survival and decreases infections in these patients. To determine the molecular mechanisms, the aim of the present study was to establish a stress model using primary human hepatocytes (PHHs) and to study the effects of insulin on the hepatic inflammatory signaling cascade. Liver tissue was obtained from general surgical patients, and PHHs were isolated and maintained in culture. Primary hepatocyte cultures were challenged with various doses of lipopolysaccharide (LPS), and the inflammatory signal transcription cascade was determined by real-time PCR. In subsequent experiments, primary hepatocyte cultures were challenged with LPS and insulin was added in various doses. Glucose was determined by colorimetric assays. PHHs treated with 100 microg/mL LPS showed a profound inflammatory reaction with increased expression of interleukin (IL)-6, IL-10, IL-1beta, tumor necrosis factor (TNF), and signal transducer and activator of transcription 5 (STAT-5). Insulin at 10 IU/mL significantly decreased IL-6, TNF, and IL-1beta at pretranslational levels, an effect associated with decreased STAT-5 mRNA expression (P0.05). Glucose concentration and cellular metabolic activity were not different between controls and insulin-treated cells. Based on our results, we suggest that primary hepatocyte cultures can be used to study the effect of LPS on the inflammatory cascade. Insulin decreases hepatic cytokine expression, which is associated with decreased STAT-5 expression.

The absorption, fluorescence and fluorescence excitation spectra for 3,20-di(tert-butyl)-2,2,21,21-tetramethyl-all-trans-3,5,7,9,11,13,15,17,19-docosanonaen (ttbP9) in dilute solutions of 2-methylbutane were recorded at temperatures over the range 120-280 K. The high photostability of this nonaene allows us to assert that it exhibits a single fluorescence and that this can be unequivocally assigned to emission from its 1(1)B(u) excited state, it being the first excited electronic state. Available photophysical data for this polyene and the wealth of information reported for shorter all-trans polyenes allow us to conclude that if the first excited electronic state for the chromophore possessed 2(1)A(g) symmetry, then the energy of such a state might have been so close to that of the 1(1)B(u) state that: 1) the radiationless internal conversion mechanism would preclude the observation of the emission from the 1(1)B(u) state reported in this work and 2) the 2(1)A(g) state reached through internal conversion would be vibrationally coupled to 1(1)B(u) and would facilitate the detection of the emission from 2(1)A(g), which was not observed in any of the solvents used in this work. The spectroscopic and photochemical implications of these findings for other polyenes are discussed.

Rapid healing of acute wounds, e.g., in burned patients, can be essential for survival. Oxidized regenerated cellulose/collagen (ORC/collagen) has been shown to improve wound healing of chronic wounds. The aim of the present study was to determine the effect of ORC/collagen on dermal and epidermal healing as well as growth factor concentration in acute wounds. Rats received a full-thickness excision wound and were treated with either ORC/collagen plus a hydrocolloid dressing or a hydrocolloid dressing alone. Planimetry, immunological assays, histological and immunohistochemical techniques were used to determine dermal and epidermal regeneration, protein concentration, and growth factor concentration. In addition, dermal vascularization and structure were determined. Wounds treated with ORC/collagen showed a significantly faster reepithelization than those treated with hydrocolloid alone, p < 0.05. This accelerated wound healing rate may be explained by significantly higher levels of platelet-derived growth factor, keratinocyte growth factor, insulin-like growth factor-I, and insulin-like growth factor binding protein-3 in the ORC/collagen group leading to antiapoptotic effects of skin cells, p < 0.05. There were no significant differences in collagen morphology or deposition, neo-angiogenesis, or vascular endothelial growth factor concentration between both treatment groups. We conclude that ORC/collagen matrix accelerates epidermal regeneration and locally increases growth factor concentrations. Increased reepithelization was associated with decreased skin cell apoptosis. Based on our data we hypothesize that the ORC/collagen matrix may also have beneficial effects on acute wounds in a clinical setting.

The careful assessment of microchimerism is essential to investigate the effects of donor bone marrow-derived cells in transplantation. We have developed a protocol to assess microchimerism based on the HLA mismatch between the recipient and the donor. Our approach combines real-time polymerase chain reaction (PCR) with sequence-specific primer PCR (SSP-PCR) to selectively amplify and measure the abundance of donor HLA alleles in DNA samples extracted from the recipient after transplant. To optimize and validate the reliability of this method at different levels of microchimerism, we tested serial dilutions of donor DNA into recipient DNA. We demonstrate that donor alleles can be readily detected and reliably measured at concentrations as low as 0.1%. This method is simple and rapid and could find practical application in the assessment of microchimerism in patients receiving organ or cellular transplants in conjunction with donor bone marrow cells infusion.

Starting from the readily available α,β-unsaturated ketone, 3-tert-butyl-4,4-dimethyl-2-pentenal, higher vinylogues, and fully terminally tert-butylated polyolefins with up to 13 consecutive conjugated double bonds have been prepared by either McMurry dimerization or Wittig chain-elongation routes. The highly unsaturated conjugated π systems, which show a remarkable stability, have been characterized by spectroscopic methods and, in many cases, by X-ray structural analysis. The yields are high enough to allow for thorough chemical reactivity studies.

Oncogenic signaling through activation of epidermal growth factor receptor (EGFR), HER-2, and hypoxia inducible-factor-1α (HIF-1α) has been implicated in gastric cancer growth and angiogenesis through up-regulation of vascular endothelial growth factor (VEGF). Recently, heat shock protein 90 (Hsp90) has been identified as a critical regulator of oncogenic protein stability, including EGFR, HER-2, and HIF-1α. We hypothesized that inhibition of Hsp90 impairs EGF- and hypoxia-mediated angiogenic signaling in gastric cancer cells and consequently inhibits angiogenesis and tumor growth. In vitro, the geldanamycin derivate 17-allylamino-17-demethoxygeldanamycin (17-AAG) led to marked reduction in constitutive and inducible activation of extracellular signal-regulated kinase 1/2, Akt, and signal transducer and activator of transcription 3 and decreased nuclear HIF-1α protein. In addition, EGFR and HER-2 were down-regulated after Hsp90 inhibition. With respect to regulation of angiogenic molecules, 17-AAG significantly reduced EGF-mediated VEGF secretion. Phosphorylation of focal adhesion kinase and paxillin were both abrogated by 17-AAG, which resulted in significant impairment of cancer cell motility. Interestingly, cytotoxic effects of 17-AAG in vitro were higher on cancer cells and gastric fibroblasts than on pericytes. In vivo, the water-soluble compound 17-dimethylaminoethylamino-17-demethoxygeldanamycin (17-DMAG; 25 mg/kg, thrice per week) significantly reduced s.c. xenografted tumor growth. By immunohistochemistry, 17-DMAG significantly reduced vessel area and numbers of proliferating tumor cells in sections. Furthermore, similar significant growth-inhibitory effects of 17-DMAG were achieved when administered as low-dose therapy (5 mg/kg, thrice per week). In conclusion, blocking Hsp90 disrupts multiple proangiogenic signaling pathways in gastric cancer cells and inhibits xenografted tumor growth in vivo. Hence, gastric cancer harbors attractive molecular targets for therapy with Hsp90 inhibitors, which could lead to improved efficacy of antineoplastic therapy regimens. [Mol Cancer Ther 2007;6(3):1123–32]

Liposomal gene transfer is an effective therapeutic approach to improve dermal and epidermal regeneration. The purpose of the present study was to define whether the biological or chemical structure of a liposome influences cellular and biological regeneration in the skin, and to determine by which mechanisms possible changes occur. Rats were inflicted a full-excision acute wound and divided into three groups to receive weekly subcutaneous injections of DMRIE liposomes plus the Lac Z gene, or DOTAP/Chol liposomes plus the Lac Z gene, or saline. Planimetry, immunological assays, histological and immunohistochemical techniques were used to determine cellular responses after gene transfer, protein expression, dermal and epidermal regeneration. DOTAP/Chol increased IGF-I and KGF protein concentration and caused concomitant cellular responses, for example, by increasing IGFBP-3, P

In the PI3K/Akt signaling pathway, »mammalian target of rapamycin« (mTOR) is a central mediator for transcription and translation of different target genes. Interestingly, the transcription factor »hypoxia inducible factor-1a« (HIF-1α) which is involved in tumor progression and metastases, is regulated by mTOR. We hypothesized that blockade of mTOR with rapamycin (RAPA) would significantly inhibit HIF-1α expression in human gastric cancer cells in vitro and impair angiogenesis and tumor growth in vivo. Effects of RAPA (10 ng/ml) on nuclear HIF-1α expression in human gastric cancer cells (TMK-1) were investigated by chemical hypoxia with desferroxamine (DFX 100 µM) and subsequently Western blot analyses. Cytotoxic effects of RAPA on TMK-1 cells were assessed by MTT assays under the same conditions. Effects of mTOR inhibition on cancer cell migration and invasion were tested in in vitro assays using modified Boyden chambers. Modulation of angiogenesis in vivo by RAPA treatment (1.5 mg/kg/day) (or vehicle) was first investigated in the dorsal skinfold chamber (DSFC) model in athymic nude mice (n = 6/group). Effects of RAPA on tumor growth were subsequently investigated in a subcutaneous tumor model. Tumor diameters were measured every second day and volumes calculated. mTOR blockade led to a 70% reduction in HIF-1α expression in vitro. MTT assays revealed modest cytotoxic effects of RAPA. In contrast, cell migration and invasion were significantly inhibited (P < 0.05). In vivo, in the DSFC model, RAPA treatment significantly reduced angiogenesis and microvessel density (P < 0.05), compared to controls. In addition, in the SQ model, RAPA led to significant reduction of tumor volumes (P < 0.05) and tumor weights (P < 0.05). In conclusion, mTOR inhibition with rapamycin could be a promising approach to augment antineoplastic/antiangiogentic therapy regimes for the treatment of gastric cancer.

Growth factors have been shown to modulate the complex cascade of wound healing, however, interaction between different growth factors during dermal and epidermal regeneration is still not entirely defined. We have recently shown that exogenous liposomal gene transfer of cDNA results in physiologic expression and response in an acute wound. In the present study we determined the interaction between insulin-like growth factor-I (IGF-I), a mesenchymal growth factor, administered as liposomal cDNA, with other dermal and epidermal growth factors on collagen synthesis in an acute wound. Sprague-Dawley rats were given a scald burn to inflict an acute wound and divided into two groups to receive weekly subcutaneous injections of liposomes plus a beta-galactosidase containing plasmid (Lac Z [0.2 microg, vehicle]), or liposomes plus the IGF-I cDNA containing plasmid (2.2 microg) and Lac Z (0.2 microg). Immunological assays, histological and immunohistochemical techniques were used to determine growth factor concentration and different types of collagen (I, III, and IV) after IGF-I cDNA gene transfer. IGF-I cDNA transfer accelerated reepithelization and was associated with increased levels of IGF-I, fibroblast growth factor, keratinocyte growth factor, vascular endothelial cell growth factor, and platelet-derived growth factor protein expression. IGF-I cDNA had no effect on transforming growth factor-beta. IGF-I cDNA significantly increased type IV collagen while it had no effect on types I and III collagen. Exogenously administered IGF-I cDNA increased protein concentrations of keratinocyte growth factor, fibroblast growth factor, platelet-derived growth factor, and type IV collagen. We conclude that liposomal IGF-I gene transfer can accelerate wound healing without causing an increase in types I and III collagen expression.

Proteins are primary targets in drug discovery. However, with a few rare exceptions, they are unable to cross cell membranes, a limitation that prevents the full exploitation of their therapeutic potential. Major advances have been recently made through a novel approach of protein and peptide delivery into cells known as protein transduction or protein therapy. Proteins and peptides can be directly transferred to cells when covalently linked to protein transduction domains (PTD), small peptides that can freely cross cell membranes with low lytic activity (1–3). The mechanism of cellular translocation of PTD are currently poorly understood. Most of the PTD described in the literature have a high content of basic residues. It is believed that the interaction with the negative cell membrane environment has an important role in the translocation process, and the mechanism of cell internalization may differ for each of the PTD. Several PTD have been identified in naturally occurring proteins. The most commonly studied are homeodomain transcription factors such as antennapedia (4), the herpes simplex virus type 1 protein VP22 (5), and the human immunodeficiency virus (HIV) transactivator TAT protein ( 6– 7). In addition, a new gamut of peptides with PTD capabilities have been recently identified. Some of these new peptides are derived from natural proteins, whereas others are synthetic peptides. The PTD included in these groups are described below, with emphasis on the TAT-PTD and its potential application in organ and cell transplantation.

Liposomal gene transfer is an effective therapeutic approach for the treatment of several pathophysiologic states. The purpose of the present study was to define whether gene transfer of multiple genes is a feasible approach and whether this approach would be more effective than the single transfer of cDNA. Rats were inflicted an acute wound and divided into four groups to receive weekly subcutaneous injections of liposomes plus the Lac-Z gene (0.22 microg, vehicle), or liposomes plus the insulin like-growth factor-I (IGF-I)cDNA (2.2 microg) and Lac Z gene (0.22 microg), or liposomes plus the keratinocyte growth factor (KGF) cDNA (2.2 microg) and Lac Z gene (0.22 microg), or liposomes plus the IGF-I/KGF cDNA (2.2 microg) and Lac Z gene (0.22 microg). Planimetry, immunological assays, histological and immunohistochemical techniques were used to determine molecular mechanisms after gene transfer, protein expression, dermal and epidermal regeneration. IGF-I/KGF cDNA transfer increased IGF-I and KGF protein concentration and caused concomitant cellular responses, for example,by increasing IGFBP-3, P

The electronic structures of the a,a,w,w-tetra-t-Bu substituted conjugated polyenes have been investigated by UPS and quantum chem. calcns. The all-trans-hexatriene (1), octatetraene (2), decapentaene (3), and the tetradecaheptaene (4) have essentially planar polyene chromophores and accordingly their p MOs are spread by about 3-5 eV. On the other hand, interaction of the double bonds is limited in the moderately twisted 1,1,6,6-tetra-tert-butyl-cis-hexatriene (5) and the highly distorted cis-1,1,6,6-tetra-tert-butyl-3,4-dimethylhexatriene (6). The first UV-Vis absorption of a,a,w,w-tetra-t-butyl-polyenes with three to thirteen conjugated C=C double bonds is linearly correlated with the PM3 calcd. HOMO-LUMO energy gap.

INTRODUCTION. Microsomal Heme Oxygenase-1 (HO-1) has been identified as a ubiquitous stress protein induced in many cell types by various stimulants such as hemolysis, inflammatory agents, oxidative stress, heat shock, and growth factors[1]. HO-1 is the enzyme that controls the degradation pathway of heme by catalyzing its oxidation into biliverdin, carbon monoxide, and iron. Although the role of HO-1 induction in oxidative stress is not completely understood, it has been shown that induction of HO-1 expression results in protection from cytokine-induced apoptosis and oxidative stress in in vitro cell culture and in various animal models, including pancreatic islet transplantation[2]. Thus, expression of HO-1 through gene therapy protocols might prove useful to reduce the deleterious effects of oxidative stress in islet isolation as well as to prevent damage in the peri-transplant period. However, viral vectors or other transfection methods available for transduction of genes into islets have limited efficacy and the presence of viral antigens (especially adenovirus) may potentially induce immunological responses against the transfected islets. Moreover, the long-term effects of genetic manipulations of islets, in particular those affecting apoptosis, may have undesirable long-term effects such as impaired mitochondrial signals regulating insulin secretion. Proteins can be directly transferred to cells when they are linked to protein transduction domains (PTDs), small peptide domains that can freely cross cell membranes[3]. In particular, proteins fused to an 11-amino acid protein transduction domain (PTD) from the human immunodeficiency virus transactivator of transcription (TAT) protein, readily transduce many cell types including pancreatic islets[4]. In this study we have characterized a functional HO-1 protein fused to a PTD METHODS. Cloning and Related Techniques. The recombinant TAT antiapoptotic fusion proteins were generated by subcloning the coding region of the murine HO-1 gene in frame with the TAT leader peptide (amino acids 47-58 YGRKKRRQRRR) in a bacterial expression generously provided by Steven Dowdy from Washington University School of Medicine, St Louis. A 6xHis-affinity tag allowed the purification of the fusion protein through affinity chromatography.

The all-trans pentaene, 3,12-di(tert-butyl)-2,2,13,13-tetramethyl-3,5,7,9,11-tetradecapentaene (ttbP5) fluoresces in two different regions of the visible spectrum. It produces an extremely weak emission in the gas phase that can also be detected in the condensed phase; such an emission exhibits a negligible Stokes shift with respect to the 1Ag-->1Bu absorption transition and can in principle be assigned to the 1Bu-->1Ag emission of the compound. ttbP5 also exhibits a second fluorescence emission at approximately 520 nm in both the gas phase and the condensed phase. The emission in the condensed phase increases in strength and structure, with no change in spectral position, as the solvent viscosity increases by effect of the solution temperature being lowered. The spectral behavior of this pentaene (ttbP5) is different enough from that reported [J. Catalan et al., J. Chem. Phys. 128, 104504 (2008)] for its tetraene counterpart (ttbP4) to warrant a separate analysis in order to facilitate a better understanding of the way the photophysics of these polyenes changes as their chain is lengthened.

INTRODUCTION. With the development of new strategies to avoid immunological rejection, transplantation of pancreatic islets has become a therapeutic reality to cure diabetes (1-2). However, despite the progress in islet isolation procedures, a single donor transplant does not provide enough islets to attain insulin independence. There is evidence that significant loss of islet cells takes place during isolation due to the triggering of apoptosis (3). There is also substantial evidence that reduction of isolation-induced apoptosis can improve the success rates of islet transplantation. The goal of this proposal is to address the needs for reduced apoptosis and improved viability of islets in conjunction with islet isolation procedures. We present in this study novel transduction methods that allow manipulation of islets to reduce apoptosis. Proteins can be directly transferred to cells when they are linked to protein transduction domains (PTDs), small peptide domains that can freely cross cell membranes. In particular, proteins fused to an 11-amino acid PTD from the human immunodeficiency virus, the TAT protein, readily diffuse across membranes and are efficiently transduced into virtually any cell type (4). The expression as well as the biological function of the TAT fusion protein are temporary without permanent modification of the cell sensitivity to apoptosis, thus avoiding undesirable long-term effects.

In 1947, McCrea [1], as well as Burner and Stone [2], observed that when a filtrate from a culture of Clostridium was added to a suspension of mammalian erythrocytes, the erythrocytes lost their ability to be agglutinated by the influenza virus. The filtrate apparently contained an agent that destroyed the erythrocyte receptor for the virus, and that appeared to have enzyme-like characteristics. Burnet and Stone also pointed out that the agent was similar to a substance present in the influenza virus particles which also destroyed the ability of erythrocytes to be agglutinated by other viruses [3]. Subsequent studies revealed that the substance in the influenza virus liberated enzymatically a compound identified as sialic acid from brain mucolipids [4]. In retrospect, the liberated compound proved to be identical to a low molecular mass substance released from ovomucin by the influenza virus and described by Gottschalk and Lind [5] 7 yr earlier. The enzyme responsible for the release of sialic acid was denoted sialidase by Heimer and Meyer [6] and neuraminidase by Gottschalk [7]. Both terms are now used indiscriminately, although some authors point out that the product of the enzymatic reaction is really sialic acid and not neuraminic acid (sialic acid is the free Nor N,0-substituted derivative of neuraminic acid). Later, studies carried out in several laboratories demonstrated that neuraminidase is present not only in viruses (mostly orthoand paramyxoviruses) and bacteria (in particular in Eubacteriales and Pseudomonadales), but also in mammalian and avian tissues (reviewed in [8]).

Delivering cytoprotective proteins/peptides into pancreata prior to islet isolation through protein transduction (PT) is a novel strategy to enhance the yield of viable transplantable islets. Previous work has shown that the protein transduction domain PTD-5 efficiently transduced islets via the pancreatic duct. TAT/PTD is a well-characterized PTD with the capability to cross even the hemato–encephalic barrier. In this study, we investigated the utilization of the 11-aa TAT protein transduction domain (TAT/PTD) to deliver peptides or proteins of different sizes ranging from 1.2 to 120 kDa, as the TAT/PTD and TAT/PTD-BH4 peptide, or the TAT/PTD–β-galactosidase fusion protein, into islets through the pancreatic duct. Using flow cytometry analysis we found that TAT/PTD derivatives transduced practically 100% of the islet cell population. Moreover, confocal laser scanning microscopy in live, nonfixed islets confirmed these results assessing transduction of TAT/PTD molecules into intact nondisaggregated islets. TAT–β-galactosidase peptide conjugated to FITC was not compartment selective, as both cytoplasmic and nucleic cellular compartments were positively stained. Furthermore, TAT–β-galactosidase peptide delivery was highly effective, as even cells located in the inner core region of the islets were transduced. Finally, transduced TAT–β-galactosidase fusion protein was biologically active after islet isolation and manipulation, and islet insulin secretion capability was not compromised by peptide transduction. These findings suggest that the transduction of chimeric TAT/PTD proteins can represent an efficient tool of molecular delivery independent of the size, to enhance or modify a specific phenotype at the nuclei or cytoplasmic level.