bluuebunny/crossref_metadata_2025 huggingface dataset:
ABO‐incompatible allotransplantation as a basis for clinical xenotransplantation
Authors: Stussi Georg , West Lori , Cooper David K. C. , Seebach Jörg D.
Published: August 2006

Abstract:  The 8th Congress of the International Xenotransplantation Association (IXA), held in Goteborg in September, 2005, immediately followed the 2nd International Symposium on ABO‐Incompatibility in Transplantation, both congresses organized by Michael Breimer and Lennart Rydberg. The Proceedings of the Symposium on ABO‐Incompatibility in Transplantation have been published (Xenotransplantation 2006; 13 (2)). The present paper provides an overview of a workshop held at the 8th Congress of the IXA, and highlights the immunological concepts emerging from ABO‐incompatible allotransplants and discusses them in relation to xenotransplantation.

Using specified immunomodulatory protocols, ABO‐incompatible solid organ allotransplantation has become a clinical reality for a small number of patients over the last two decades. ABO‐incompatible adult kidney and infant heart transplants have similar patient and graft survivals as their ABO‐compatible counterparts. In contrast, ABO‐incompatibility is present in up to 30% of all patients receiving hematopoietic stem cell transplants in the absence of specific immunomodulation, without affecting overall survival. Consequently, ABO‐incompatible solid organ transplants and hematopoietic stem cell transplants may serve as in vivo models to elucidate the immunological mechanisms of accommodation and/or tolerance in the clinical setting. Because of similarities in the immunological hurdles that need to be overcome, knowledge obtained from ABO‐incompatible allotransplantation might further promote advances in the field of xenotransplantation. The similarities and differences between ABO‐incompatible allotransplantation and xenotransplantation are discussed.
NK‐cell‐dependent acute xenograft rejection in the mouse heart‐to‐rat model
Authors: Chen Daxin , Weber Michele , Lechler Robert , Dorling Anthony
Published: August 2006

Abstract:  Background:  Acute humoral xenograft rejection is characterized by widespread intravascular thrombosis with a significant NK‐cell and macrophage infiltrate. Although in vitro and ex vivo data have shown that NK cells are capable of killing xenogeneic tissue, the precise role they play in vivo is still not certain. Consequently, there are few tested strategies for dealing with NK‐cell‐mediated rejection, should this prove to be a problem. One reason for this has been the lack of a relevant rodent model in which rejection by these cells can be easily studied.

Methods:  Prior to transplantation of mouse hearts, we depleted rat recipients of fibrinogen using a snake venom, ANCROD, from the Malayan pit viper. Graft survival was examined by manual palpation and the rejected hearts were examined by histology. Levels of circulating interferon gamma (IFN‐γ), used as a surrogate marker for NK‐cell activation, were determined by an enzyme‐linked immunosorbent assay.

Results:  Depletion of fibrinogen to approximately 5% of normal allowed surgery without a significant increase in the technical failure rates and prolonged graft survival compared with that seen in unmanipulated rats. Rejected hearts showed no evidence of intravascular thrombosis but did show significant antibody and complement deposition. There was little T‐cell infiltration and cyclosporin had no influence on survival. Instead, hearts were infiltrated with NK cells and macrophages and rejection was associated with significant IFN‐γ production. Depletion of NK cells with anti‐asialo‐GM‐1 from ANCROD‐treated recipients led to a further significant prolongation of graft survival.

Conclusions:  Inhibition of intravascular thrombosis by fibrinogen depletion, in the absence of any other manipulation, unmasks NK‐cell‐dependent acute xenograft rejection in the mouse‐to‐rat heart transplantation model. This relatively simple model is expected to be useful to investigate the mechanisms of NK‐cell‐mediated rejection and to provide insight into the types of graft manipulation that could modify this process.
Xenotransplantation of purified pre‐natal porcine beta cells in mice normalizes diabetes when a short anti‐CD4–CD8 antibody treatment is combined with transient insulin injections
Authors: Veld Peter In't , Pavlovic Dejan , Bogdani Marika , Pipeleers‐Marichal Miriam , Pipeleers Daniel
Published: August 2006

Background:  Pre‐natal porcine endocrine islet cell grafts were recently shown to contain immature beta cells with a marked potential for growth and differentiation following transplantation, and hence for a progressive and long‐term correction of diabetes in immune‐incompetent mice. The present study investigates whether these grafts are also capable of correcting hyperglycemia in immune‐competent mice receiving a short treatment with anti‐CD4–CD8 antibodies.

Methods:  Pure endocrine islet cell grafts with 0.5 to 1.0 million beta cells were prepared from pre‐natal pigs and transplanted under the kidney capsule of alloxan‐diabetic CBA/Ca mice. Survival, growth and function of implanted beta cells were followed by measuring plasma porcine C‐peptide and glucose, and graft insulin content at start and at post‐transplant (PT) week 35. The effect was studied of a 5‐day treatment with non‐depleting anti‐CD4 YTS177 and depleting anti‐CD8 YTS169 antibody, either without or with transient insulin injections.

Results:  Without antibody treatment, all graft recipients remained porcine C‐peptide negative and died. Antibody treatment decreased CD4‐expression and percentage CD8 cells for 10 and 18 weeks respectively. It resulted in a 30 week‐survival of nine out of 14 graft recipients; all nine had progressively become C‐peptide positive but only one proceeded to normoglycemia. When antibody treatment was combined with transient insulin injections, 11 out of 14 graft recipients survived long‐term, eight became C‐peptide positive and six were normoglycemic at PT week 30. In both groups, surviving recipients exhibited a graft insulin content that was 6‐ to 9‐fold higher than at implantation.

Conclusions:  Pre‐natal porcine beta cells grow and differentiate when transplanted in diabetic immune‐competent mice that have been transiently immune suppressed with anti‐CD4 and anti‐CD8 monoclonal antibodies. They develop metabolic control when recipients are also transiently treated with insulin injections.
Compatibility of porcine and human interleukin 2: implications for xenotransplantation
Authors: Zhang Baojun , Ge Basheng , Xia Xuepei , Liu Jun , Sun Liguang , Rao Enyu , Zhao Yong
Published: August 2006

Abstract:  Background:  Xenotransplantation provides a possible solution to the severe shortage of allogeneic organ donors. The pig, which shares many physiological similarities with humans, makes it an optimal species for preclinical experimentation and clinical applications. Interleukin 2 (IL2) is a potent growth factor secreted primarily by T helper lymphocytes and it is vital to the cellular expansion required for a productive immune response and the development and peripheral expansion of CD4+CD25+ regulatory T cells. Therefore, it is essential to understand of the compatibility of IL2 between pigs and humans.

Methods:  We first compared the cDNA and protein sequences and the crystal structures of human and porcine IL2 and IL2 receptors, respectively. The effect of IL2 to induce T cell proliferation was determined by 3H‐thymidine incorporation and cell cycle detection.

Results:  Porcine IL2 induced very limited proliferation of human lymphocytes while it functioned well on porcine lymphocytes. Human IL2 had remarkably reduced effects on porcine lymphocytes whereas it worked well on human lymphocytes.

Conclusion:  Our present study showed that the interaction of IL2 and IL2R across species might have defects. Together with the wide physiological functions of IL2, our data indicated that physiological disorders could be caused by the poor function of xenogeneic donor IL2 on host cells in full hematopoietic chimera. Our data suggested an additional potential advantage for the mixed xenogeneic chimeras.
Effect of tandem forms of DAF(CD55) on complement‐mediated xenogeneic cell lysis
Authors: Miyagawa Shuji , Fukuta Daisuke , Kitano Etsuko , Kobayashi Chizuko , Fumimoto Yuichi , Shirasu Akio , Hattori Hiroyuki , Shirakura Ryota , Fukuzawa Masahiro
Published: August 2006

Abstract:  Background:  It is difficult to produce a transgenic animal with high expression of decay‐accelerating factor (CD55: DAF) or other molecules. The purpose of this study was to assess the effect of tandem forms of DAF on a xenogeneic cell membrane against human complement.

Methods:  cDNAs of the delta‐Short Consensus Repeat (SCR) 1‐DAF, the double‐DAF, the triple‐DAF, and the tetra‐DAF with a FLAG‐tag were established. Chinese hamster ovary (CHO) cell lines and a pig endothelial cell (PEC) line expressing these molecules were established. The amelioration of complement‐mediated lysis by the transfectant molecules on these cells was examined. The CHO cell transfectants were also incubated with normal human serum, and the amount of C3 deposited was determined by FACS analysis.

Results:  Stable CHO cells and PEC transfectants, in which each molecule was clearly expressed, and Western blots showed that each band corresponded to the expected molecular weight. The extent of amelioration of complement‐mediated lysis by these four molecules was then examined. A clear tendency was found, as follows: The higher the tandem number of DAF, the greater was the effect on cytotoxicity. Additional experiments focusing on triple‐DAF and tetra‐DAF did not indicate any significant difference in complement‐mediated lysis. Consistent with the complement‐regulatory ability, the inhibitory effect of the deposition of C3 fragments by these molecules was closely related to the degree of amelioration.

Conclusion:  These data indicate that tandem DAF, especially a triple‐DAF, is a very effective form for protecting against complement activation.
Recognition of a carbohydrate xenoepitope by human NKRP1A (CD161)
Authors: Christiansen Dale , Mouhtouris Effie , Milland Julie , Zingoni Alessandra , Santoni Angela , Sandrin Mauro S.
Published: August 2006

Abstract:  Background:  Many immunologically important interactions are mediated by leukocyte recognition of carbohydrates via cell surface receptors. Uncharacterized receptors on human natural killer (NK) cells interact with ligands containing the terminal Galα(1,3)Gal xenoepitope. The aim of this work was to isolate and characterize carbohydrate binding proteins from NK cells that bind αGal or other potential xenoepitopes, such as N‐acetyllactosamine (NAcLac), created by the deletion of α1,3galactosyltransferase (GT) in animals.

Methods and results:  Initial analysis suggested the human C‐type lectin NKRP1A bound to a pool of glycoconjugates, the majority of which contained the terminal Galα(1,3)Gal epitope. This was confirmed by high level binding of cells expressing NKRP1A to mouse laminin, which contains a large number of N‐linked oligosaccharides with the Galα(1,3)Gal structure. The consequence of removing the terminal αGal was then investigated. Elevated NAcLac levels were observed on thymocytes from GT−/− mice. Exposing NAcLac on laminin, by α‐galactosidase treatment, resulted in a significant increase in NKRP1A binding.

Conclusions:  NKRPIA binds to the αGal epitope. Moreover, exposing NAcLac by removal of αGal resulted in an increase in binding. This may be relevant in the later phases of xenotransplant rejection if GT−/− pigs, like GT−/− mice, display increased NAcLac expression.
A study of the xenoantigenicity of neonatal porcine islet‐like cell clusters (NPCC) and the efficiency of adenovirus‐mediated DAF (CD55) expression
Authors: Omori Takeshi , Nishida Toshirou , Komoda Hiroshi , Fumimoto Yuichi , Ito Toshinori , Sawa Yoshiki , Gao Congxiao , Nakatsu Shino , Shirakura Ryota , Miyagawa Shuji
Published: August 2006

Abstract:  Background:  The pig pancreas is considered to be the most suitable source of islets for xenotransplantation in patients with type I diabetes. The objective of this study was to assess the antigenicity of neonatal porcine islet‐like cell clusters (NPCC), including the Galα1–3Galβ1–4GlcNAc‐R (α‐Gal) and Hanganutziu–Deicher (H–D) antigens, and the pathway involved in human complement activation. The efficiency of expression of human decay‐accelerating factor (DAF: CD55) on NPCC by adenoviral transduction was also examined, and the functional capacity of DAF was also estimated.

Methods:  The deposition of human natural antibodies, immunoglobulin (Ig)G and IgM, and the expression of α‐Gal and H–D antigens on NPCC were investigated by FACS analysis. The downregulation in the antigenicity to human natural antibodies, including the α‐Gal and H–D antigens on NPCC by treatment with tunicamycin, PDMP and neuraminidase were also examined. In addition, complement‐mediated islet lysis was examined using factor D‐deficient and C1‐deficient sera. An adenovirus encoding DAF under the control of the cytomegalovirus promoter, Ad.pCMV‐DAF, was then constructed, and used for transducing NPCC. The amelioration of complement‐dependent cytotoxicity of the NPCC by the transduced DAF was assessed as an in vitro hyperacute rejection model of a pig to human xenograft.

Results:  The NPCC clearly expressed the α‐Gal epitope, and the human natural antibodies, IgG and IgM, and the anti‐H–D antibody also reacted with the NPCC. Treatment of NPCC with tunicamycin led to a drastic reduction in the extent of deposition of IgG, indicating the importance of N‐linked sugars on the islets, presumably related to α‐Gal expression on N‐linked sugars. Neuraminidase treatment indicated the presence of, not only the H–D antigen, but also other sialic acid antigens which reacted with the human natural antibody, especially IgG. The complement deposition of factor B on NPCC was clear, and the alternative pathway‐mediated NPCC killing accounted for approximately 30% of that by the total complement pathway. On the other hand, approximately 90% of the NPCC could be transduced to express DAF by the adenovector, Ad.pCMV‐DAF. The expressed DAF showed an approximately 50–62% suppression in complement‐dependent NPCC lysis.

Conclusion:  The origin of the antigenicity of NPCC is mainly N‐linked sugars including α‐Gal and sialic acid antigens, and NPCC expressed the transduced molecule in high efficiency by the adenovector.
The Effect of Nitric Oxide Inhibition and Temporal Expression Patterns of the mRNA and Protein Products of Nitric Oxide Synthase Genes During In Vitro Development of Bovine Pre‐implantation Embryos
Authors: Tesfaye D , Kadanga A , Rings F , Bauch K , Jennen D , Nganvongpanit K , Hölker M , Tholen E , Ponsuksili S , Wimmers K , Montag M , Gilles M , Kirfel G , Herzog V , Schellander K
Published: August 2006

Contents

This study was conducted to determine the effect of Nitric oxide (NO) inhibition in bovine in vitro development and expression analysis of the three Nitric oxide synthase (NOS) isoforms: endothelial (eNOS), neuronal (nNOS) and inducible (iNOS), mRNA and protein in bovine oocytes and embryos. Selective inhibitor of NOS, N‐ω‐nitro‐l‐arginine methyl ester (l‐NAME) was applied at different doses (0, 0.1, 1 and 10 mm) in maturation (experiment 1A), culture medium (experiment 1B) and in both maturation and culture media (experiment 1C). No significant differences were observed in cleavage and blastocyst rates when oocytes were matured in the presence of l‐NAME as long as the inhibitor was omitted during fertilization and culture. However, significantly lower blastocyst rates were observed when l‐NAME was present at higher level (10 mm) in culture medium alone and in both maturation and culture media. In experiment 2, mRNA isolated from triplicate pools of oocytes and embryos (n = 15–20) was subjected to quantitative real time reverse transcription polymerase chain reaction to investigate the expression of eNOS, iNOS and nNOS mRNA in normal IVP bovine oocytes and embryos. While eNOS and iNOS transcripts were detected at higher level in oocytes (immature and mature), two‐cell and four‐cell stage embryos, the nNOS was detected only in immature oocyte, two‐cell and morula stages. In experiment 3, eNOS and iNOS protein expression analysis was performed in IVP oocytes and embryos and both proteins were detected in the cytoplasm and the nuclei (weak) of oocytes and embryos. These data provide the first evidence for the role of NO production and the presence of mRNA and protein products of NOS isoforms during bovine embryogenesis.
The Role of Oestradiol in the Uterine Peristalsis in the Perfused Swine Uterus
Authors: Maltaris T , Dittrich R , Widjaja W , Sindhuwinata C , Hoffmann I , Beckmann MW , Mueller A
Published: August 2006

Contents

This study was designed to examine the effects of oestradiol (E2) on sperm transport in the swine uterus. The bicornuate swine uterus is optimal for the study of the uterine transport and peristalsis because the influence of various factors can be examined on each uterine horn independently. Forty swine uteri (with or without ovarectomy) were perfused for a period of up to 7 h. Two different E2 concentrations (3 or 30 pg/ml) in the perfusion medium were administered for 30 min unilaterally. Through an intracervical catheter 1 ml of a high concentrated dextran blue solution was administered directly in the upper part of the cervix. After bilateral perfusion of the swine uterus with a bolus of 0.3 IU oxytocin the distribution of coloured particles was assessed macroscopically before and after incision of the uterine horns. Coloration was evaluated by two observers blinded to the site‐specific administration of E2. In the 10 ovarectomized uteri with the 3 pg/ml E2 concentration a unilateral distribution towards the side of oestradiol administration was observed in six uteri, in four it was a bilateral distribution. In the 10 non‐ovarectomized uteri with the 3 pg/ml E2 concentration a uni‐ and ipsilateral coloration was observed in five uteri, in five it was a bilateral distribution. In the 20 uteri with 30 pg/ml E2, a unilateral coloration of the uterus horns was observed in all uteri. Oestradiol is one of the main factors, which influences the direction of the sperm transport in a dose‐dependent manner, in the perfused swine uterus.
Transcript Profiles of Some Developmentally Important Genes Detected in Bovine Oocytes and In Vitro‐produced Blastocysts Using RNA Amplification and cDNA Microarrays
Authors: Mamo S , Sargent CA , Affara NA , Tesfaye D , El‐Halawany N , Wimmers K , Gilles M , Schellander K , Ponsuksili S
Published: August 2006

Contents

To study the mRNA transcript profiles of some potential candidate developmental genes during bovine oocyte and blastocyst stages, RNA amplification procedures, cDNA microarray of 82 target genes spotted onto glass slide and real‐time polymerase chain reaction (PCR) were used. Messenger RNAs were isolated from in vitro‐produced bovine matured oocytes and blastocysts. Using equal amounts of input mRNAs but different cycles of amplifications, cDNAs were produced and served as template for RNA amplification by the in vitro transcriptions. After amplification, the RNA yields transcribed from cDNAs of different cycles were evaluated both by hybridization on the cDNA microarrays and by using real‐time PCR techniques. The analyses indicated best results from lower amplification cycle templates with consistent signals at hybridization. Generally, the RNA yield was directly proportional to the amplification cycle but inversely related with signal consistency at repeated hybridizations. Using the protocols established, equal amounts of amplified RNA from matured oocytes and blastocysts were hybridized to the array. Analyses of replicated hybridizations indicated that 35 transcripts were differentially expressed. Most of these were not described in previous bovine embryo studies. Independent analyses of 23 transcripts with real‐time PCR and unamplified RNA confirmed the results of 22 genes. Moreover, the functional analyses showed various roles related to development. Hence, it is possible to conclude that the genes identified here are potential candidates for characterizing developmental competence, and that the methods established can be used for large‐scale gene expression analysis with more comprehensive arrays.
The effectiveness of a health promotion programme for women in southern Taiwan
Authors: Chang Su‐Hsien , Fang Miao‐Chun , Yang Ya‐Shu
Published: August 2006

The purpose of this study was to assess the effectiveness of an 8 week aerobic exercise class for women on changes in their physical activity behaviour. This was a descriptive, retrospective and comparison study. Because the data were collected after an 8 week aerobic exercise class, there were two groups of subjects. Subjects in the exercise group had attended exercise classes, but subjects in the control group had not. Results showed statistically significant differences between the two groups in the frequency/week of moderately intense exercise‐related activities, caloric expenditure/week in all exercise‐related activities and caloric expenditure/week in moderately intense exercise‐related activities. However, there was no statistically significant difference in the frequency/week of all exercise‐related activities. The result that women in the exercise group practised physical activities more than women in the control group might be related to their high exercise self‐efficacy expectations.
Veganism and osteoporosis: A review of the current literature
Authors: Smith Annabelle M
Published: August 2006

The purpose of this review is to examine the current literature regarding calcium and Vitamin D deficiencies in vegan diets and the possible relationship to low bone mineral density and incidence for fracture. Prominent databases were searched for original research publications providing data capable of answering these questions: (i) Do vegans have lower‐than‐recommended levels of calcium/Vitamin D? (ii) Do vegans have lower bone mineral density than their non‐vegan counterparts? (iii) Are vegans at a greater risk for fractures than non‐vegans? The findings gathered consistently support the hypothesis that vegans do have lower bone mineral density than their non‐vegan counterparts. However, the evidence regarding calcium, Vitamin D and fracture incidence is inconclusive. More research is needed to definitively answer these questions and to address the effects of such deficiencies on the medical and socioeconomic aspects of life.
Baccalaureate nursing studies: voyaging towards discovery
Authors: Birks Melanie , Chapman Ysanne , Francis Karen
Published: August 2006

Postregistration nursing courses are designed to enable registered nurses to convert their existing certificate or diploma qualifications to degree level. This paper presents the findings of a preliminary exploratory, descriptive investigation into the experience of nurses undertaking postregistration degree studies in Malaysian Borneo. The unique nature of this environment presents challenges for these students and these are examined using the metaphor of a ship’s voyage. These experiences are explored within themes of setting sail, finding one’s sea legs, all in the same boat and discovering the treasure. The findings indicate that, for most of the participants, completion of a baccalaureate postregistration course was a long‐held goal. A number of challenges were faced during the course, including managing time, the stress of conflicting priorities and the adjustment to tertiary studies. Support from fellow students, faculty and family was crucial to successfully completing this course. Personal and professional changes were reported by all participants. These include an increased level of confidence and assertiveness, the development of critical thinking ability, and enhanced communication skills. Ultimately, participants reported a renewed attitude towards themselves and their profession.
A comparison of acute and long‐term health‐care personnel’s attitudes towards older adults
Authors: Gallagher Stephen , Bennett Kate M , Halford Jason CG
Published: August 2006

Considerable variations in the quality of care older adults receive may depend much on the attitude of staff towards them. The attitudes of nurses, assistant personnel and porters towards older adults were assessed. Determinants affecting this judgement, such as age, gender, education, years in practice and care setting, were also assessed. Ninety‐nine (acute) and 87 (long‐term) hospital employees completed the self‐report Kogan’s Attitude Towards Old People scale. Significant statistical differences in negative attitudes were found between assistant personnel and nurses and between porters and nurses; these non‐professionals believed that older adults were irritable, grouchy, complaining and untidy. Practice area had no influence on attitudes; attitudes were, however, significantly predicted by education levels. Findings suggest that, irrespective of setting, assistant personnel and porters possess significantly greater negative attitudes towards older adults than nursing staff. Furthermore, these negative attitudes seem to be a function of lower educational achievement. Implications for informing practice, education and policy‐makers are discussed.
Missed opportunities: Nursing insights on end‐of‐life care for haematology patients
Authors: McGrath Pam , Holewa Hamish
Published: August 2006

There is now extensive consumer research to indicate that patients with haematological malignancies are not receiving appropriate or timely referrals to the palliative system. This paper begins to explore the issue from the professional perspective by presenting findings from haematology nurses on their experience with terminal care. The nursing insights have been gathered through open‐ended interviews with a national sample of nurses with extensive experience in haematology in both public and private hospitals throughout Australia. The findings resonate with the previous consumer research in that all the acute care nurses affirmed that it is their belief, based on their professional experience, that patients from these diagnostic groups typically die in the acute ward dealing with escalating technology and invasive treatments. For some, the statements could be qualified by the satisfaction that they worked in a haematology unit, aware of the death‐denying issues, trying to address the problem. Others, caught in a ‘refractory’ subculture (i.e. a subculture with a negative perception of palliative care), outlined the factors driving the lack of integration for their specific hospital. The focus of the discussion of findings is on the latter.
Evidence‐based practice: A framework for clinical practice and research design
Authors: Leach Matthew J
Published: August 2006

The evidence‐based practice (EBP) framework emerged in the early 1970s as a means of improving clinical practice. This shift towards EBP allowed health professionals to move from a culture of delivering care based on tradition, intuition and authority, to a situation where decisions were guided and justified by the best available evidence. Despite the many advantages of EBP, many practitioners remain cautious about embracing the model. Part of this opposition is due to a misunderstanding of EBP, which this paper aims to address.
Mitochondrial glutathione: Hepatocellular survival–death switch
Authors: Garcia‐Ruiz Carmen , Fernandez‐Checa José C
Published: August 2006

Steatohepatitis represents an advanced stage of fatty liver disease that encompasses alcoholic (ASH) and non‐alcoholic steatohepatitis (NASH). The progression from steatosis to steatohepatitis is poorly understood. One of the clues to this progression is the sensitization of hepatocytes to oxidative stress and cytokine‐induced cell death. Mitochondrial glutathione (mGSH), which plays a central role in the control of mitochondrial reactive oxygen species (ROS) generation, modulates the sensitivity to cell death pathways. Mitochondrial GSH depletion due to alcohol‐mediated alteration in mitochondrial membrane dynamics underlies the susceptibility of hepatocytes from alcohol‐fed models to tumor necrosis factor (TNF), and in nutritional and genetic models of hepatic steatosis, mGSH depletion occurs due to the enrichment of mitochondria in free cholesterol, resulting in decreased mitochondrial membrane fluidity. The signaling of TNF through its membrane receptor TNFR1 from complex I to complex II is similar in hepatocytes depleted or not depleted in mGSH, yet hepatocellular susceptibility to TNF occurs if mGSH is depleted. Thus, mGSH is a critical factor in the development of steatohepatitis through sensitization of hepatocytes to inflammatory cytokines, and understanding the homeostasis of cholesterol and its trafficking to mitochondria may be of relevance in the pathophysiology of ASH and NASH.
Anti‐adipogenic regulation underlies hepatic stellate cell transdifferentiation
Authors: Tsukamoto Hidekazu , She Hongyun , Hazra Saswati , Cheng Jason , Miyahara Takeo
Published: August 2006

Cirrhosis is the most important consequence of alcoholic liver disease for which liver transplantation is the only treatment option available. Transdifferentiation of hepatic stellate cells (HSC) to myofibroblastic cells (MF) is a central event in liver fibrogenesis, and understanding molecular mechanisms that underlie this cellular event provides pivotal insights into development of new therapeutic modalities for cirrhosis. To this end, the authors proposed several years ago that transdifferentiation of quiescent HSC to MF may be causally associated with transcriptional regulation known for adipocyte–preadipocytic fibroblast dedifferentiation. In support of this notion, the authors showed that adipogenic transcription factors and their downstream adipocyte specific genes are expressed abundantly in quiescent HSC and that this expression profile is lost in HM. Further, gain‐of‐function manipulations for adipogenic transcription factors such as peroxisome proliferator‐activated receptor‐γ (PPAR‐γ) and sterol regulatory element binding protein‐1c have been shown to reverse culture‐induced MF to quiescent HSC. The authors also demonstrated that tumor necrosis factor‐α and Wnt, known mediators of anti‐adipogenesis, also suppress the activity of PPAR‐γ and contribute to HSC‐MF transdifferentiation. These results reinforce the concept of adipogenic regulation essential to the quiescent phenotype and the loss of such regulation underlying HSC‐HM transdifferentiation. They also provide insights into the molecular basis for the use of PPAR‐γ agonists, which has been advocated for treatment of liver fibrosis.
Methionine adenosyltransferase and S‐adenosylmethionine in alcoholic liver disease
Authors: Lu Shelly C , Martínez‐Chantar M Luz , Mato José M
Published: August 2006

Methionine adenosyltransferase (MAT) is an essential enzyme that catalyzes the formation of the principal methyl donor S‐adenosylmethionine (SAMe). Studies in the past decade have shown that SAMe is not only a methyl donor, but also a key metabolite that regulates hepatocyte growth, death and differentiation. Abnormalities in MAT and decreased SAMe levels occur in experimental animals and humans with alcoholic liver disease. Chronic hepatic SAMe deficiency can result in the spontaneous development of steatohepatitis and hepatocellular carcinoma. This paper reviews MAT genes and SAMe in relation to alcoholic liver disease and the molecular mechanisms by which SAMe regulates hepatocyte growth and apoptosis.
Role of matrix metalloproteinase‐2 in ethanol‐induced invasion by breast cancer cells
Authors: Luo Jia
Published: August 2006

Ethanol is a tumor promoter and may enhance the metastasis of breast cancer. However, the underlying cellular/molecular mechanisms remain unknown. Amplification of ErbB2, a receptor tyrosine kinase, is found in 20–30% of breast cancer patients. Ethanol preferably stimulates invasion by breast cancer cells over‐expressing ErbB2 in vitro. Over‐expression of ErbB2 is positively associated with elevated levels of matrix metalloproteinase‐2 (MMP‐2) and MMP‐9. Ethanol at physiologically relevant concentrations activates MMP‐2 without altering its expression level in mammary epithelial cells over‐expressing ErbB2, but not in cells expressing low levels of ErbB2. The activation is dependent on c‐jun N‐terminal kinases (JNK) and reactive oxygen species. Selective inhibitors of MMP‐2 and anti‐oxidants significantly inhibit ethanol‐stimulated cell invasion. Similarly, knocking down MMP‐2 by small interference RNA induces a partial blockage on ethanol‐promoted cell invasion. Matrix metalloproteinase‐2 is predominantly expressed in stromal fibroblasts; ethanol also activates fibroblastic MMP‐2. The conditioned medium collected from ethanol‐exposed fibroblasts dramatically stimulates the invasion of breast cancer cells. The role of MMP‐2 in ethanol‐induced tumor promotion is discussed.
Regulation of Kupffer cell activity during chronic ethanol exposure: Role of adiponectin
Authors: Park Pil‐hoon , Thakur Varsha , Pritchard Michele T , McMullen Megan R , Nagy Laura E
Published: August 2006

There is a growing appreciation that adipose tissue is a multifunctional organ. In addition to its central role in lipid storage, adipose tissue secretes a diverse group of proteins, called adipokines, involved in lipid metabolism, insulin sensitivity, angiogenesis etc. Adipocytes also secrete various inflammatory and anti‐inflammatory mediators. Adiponectin, an adipokine with potent anti‐inflammatory properties, is thought to play an important role in the regulation of inflammation. The development of alcoholic liver disease is thought to involve increased pro‐inflammatory activity, mediated in part by the activation of Kupffer cells. Chronic ethanol feeding sensitizes Kupffer cells to activation by lipopolysaccharide (LPS), leading to increased production of reactive oxygen species (ROS) and tumor necrosis factor‐α (TNF‐α). Recent studies have demonstrated a hepato‐protective effect of adiponectin in the progression of alcoholic liver disease. Herein are summarized recent data demonstrating that adiponectin treatment can normalize LPS‐stimulated ROS production and TNF‐α expression in Kupffer cells after chronic ethanol feeding. These studies suggest that the hepato‐protective activity of adiponectin is due, at least in part, to a direct anti‐inflammatory effect of adiponectin on Kupffer cells.
Unfolding new mechanisms of alcoholic liver disease in the endoplasmic reticulum
Authors: Kaplowitz Neil , Ji Cheng
Published: August 2006

Intragastric ethanol feeding in mice induces expression of unfolded protein response/endoplasmic reticulum (UPR/ER) stress response genes. The proximate cause appears to be hyperhomocysteinemia, a well‐known cause of ER stress in other contexts. Hyperhomocysteinemia appears to be due to downregulation of methionine synthase. The importance of homocysteine and ER stress in the pathogenesis of liver disease was suggested by the prevention of the alcohol‐induced changes by feeding sufficient betaine to lower homocysteine via betaine homocysteine methyl transferase. The ER stress, via CHOP, causes apoptosis and CHOP null mice exhibit no apoptosis. Alcohol‐induced ER stress can activate sterol regulatory element‐binding protein (SREBP)‐1c and SREBP‐2, which contribute to the accumulation of triglyceride and cholesterol. Hyperhomocysteinemia, ER stress and pathological changes of alcohol were minimally affected by absence of tumor necrosis factor receptor 1 (TNFR1) and the effect of betaine was also independent of TNF signaling. At present ER stress as an important factor in the pathogenesis of alcoholic liver disease is an exciting new hypothesis and ongoing research will need to further clarify its contribution. Among the issues in need of further elucidation are the role of ER stress induced by alcohol in SREBP regulation and fatty liver, as well as the precise mechanism of protection by betaine: decreased homocysteine, decreased S‐adenosylhomocysteine, or increased S‐adenosylmethionine.
Alcohol and lipid metabolism
Authors: Crabb David W , Liangpunsakul Suthat
Published: August 2006

Hepatic lipid metabolism is controlled by several master transcription factors, in particular peroxisome proliferator‐activated receptor‐α (PPAR‐α) and sterol response element binding protein‐1 (SREBP‐1). Peroxisome proliferator‐activated receptor‐α is a receptor for free fatty acids (FFA), and can activate genes involved in transport, oxidation, and export of FFA. Sterol response element binding protein‐1 is a sensor for the level of cholesterol in the liver, and is able to activate genes involved in synthesis of cholesterol and FFA. Chronic ethanol treatment of cells or animals inhibited PPAR‐α function and activated SREBP. In addition, ethanol inhibited adenosine monophosphate‐dependent protein kinase (AMPK). The AMPK controls fatty acid metabolism by inhibiting acetyl‐coenzyme A carboxylase, reducing malonyl‐coenzyme A, and thereby permitting fatty acid transport into and oxidation in the mitochondrion. Adenosine monophosphate‐dependent protein kinase was inhibited in alcohol‐treated animals and cells. The mechanisms by which ethanol affects AMPK and the transcription factors are as yet incompletely understood.
Gene–environment factors that contribute to alcoholic pancreatitis in humans
Authors: Whitcomb David C
Published: August 2006

Alcohol‐associated acute and chronic pancreatitis occur in a minority of alcohol users, suggesting that most drinkers are protected from pancreatic diseases while a subset is susceptible. Ongoing studies suggest that the pathophysiology is complex and can involve multiple genetic and environmental pathways and stochastic events. Both rat models and human genetic epidemiology studies have been used to understand susceptibility and modifying factors in humans. Rat studies suggest that different types of altered pancreatic physiology occur depending on dose, they occur rapidly and that alcohol changes the immune response to recurrent pancreatic injury. Human studies suggest that PRSS1 and SPINK1 mutation increase the pancreas’ susceptibility to alcohol‐associated pancreatitis, and that tobacco smoking, and some factors, affect disease progression.
Hepatic stellate cells and the reversal of fibrosis
Authors: Kisseleva Tatiana , Brenner David A
Published: August 2006

Hepatic fibrosis is an outcome of many chronic liver diseases, such as viral and autoimmune hepatitis, and of alcohol consumption and biliary obstruction. Prolonged liver injury results in hepatocyte damage, which triggers activation of hepatic stellate cells (HSC) and recruitment of inflammatory cells into the liver. The HSC play a critical role in fibrogenesis. They produce collagen type I and secrete pro‐fibrogenic cytokines and inhibitors of matrix‐degrading enzymes (tissue inhibitor of matrix metalloproteinase), causing the production of extracellular matrix deposition over degradation. However, many clinical and experimental studies suggest that this process can be reversed, including the apoptosis of activated HSC. Thus, HSC represent an appealing target for antifibrotic therapy. This review will focus on some aspects of etiology and molecular pathogenesis of liver fibrosis and the reversal of fibrosis.
Transcriptional regulation of stellate cell activation
Authors: Friedman Scott L
Published: August 2006

The identification of activated hepatic stellate cells and related cell types as key fibrogenic effectors during liver injury has led to intense evaluation of transcriptional events underlying their behavior. While initial studies focused on characterizing interactions between transcription factors and regulatory regions within gene promoters, epigenetic mechanisms have emerged as major determinants of gene activation and repression, in particular histone acetylation and promoter methylation, as well as other complex conditional interactions that underlie global changes in gene expression. Three examples are provided that illustrate how stellate cell activation may be controlled by widely divergent regulatory pathways, including alternative splicing of a growth inhibitory transcription factor (Kruppel‐like factor‐6), epigenetic regulation of a factor regulating stellate cell survival (nuclear factor κB), and regulation of a transcription factor whose expression maintains stellate cell quiescence (LIM homeobox gene 2 [Lhx2]). These complex cascades illustrate how clarifying the finely tuned interdependent layers of transcriptional, translational, post‐translational and epigenetic gene regulation in stellate cells is raising new prospects for therapy of hepatic fibrosis.
Matrix metalloproteinases, the pros and cons, in liver fibrosis
Authors: Han Yuan‐Ping
Published: August 2006

Residing in the space of Disse within loose extracellular matrix (ECM) resembling that in basement membranes, the hepatic stellate cells (HSC) remain in quiescence as vitamin A storage cells. In response to liver injury HSC undergo morphologic and functional trans‐differentiation, converting from vitamin A‐storing, star‐like cells into contractile myofibroblastic cells, a process called activation. Accompanying cellular activation, the ECM components in the space of Disse switch from matrices rich in type‐IV collagen and laminin, into condensed interstitial ECM, indicating that proteolytic degradation may occur to change the microenvironment in sinusoids as well as the fate of HSC. Indeed, matrix metalloproteinases (MMP), a family of ECM degradative enzymes, are promptly expressed by HSC in response to diverse hepatic toxins. In vitro experiments also demonstrated the role of MMP in activation of HSC cultured in 3‐D ECM. Conversely, MMP may also contribute to regression of liver fibrosis through cleavage of the fibrillar ECM and promotion of apoptosis among the activated HSC. Thus, MMP play dual roles both bad and good in liver fibrosis, depending on the timing.
Battle‐scarred pancreas: Role of alcohol and pancreatic stellate cells in pancreatic fibrosis
Authors: Apte Minoti V , Pirola Romano C , Wilson Jeremy S
Published: August 2006

Pancreatic stellate cells (PSC) are now recognized as the key mediators of pancreatic fibrosis, a characteristic feature of chronic pancreatitis. The role of PSC in alcoholic pancreatic fibrosis has been examined in vivo (using pancreatic tissue from patients with alcohol‐induced chronic pancreatitis and from animal models of experimental pancreatitis) and in vitro (using PSC in culture). These studies indicate that PSC are activated early in the course of pancreatic injury and are the predominant source of collagen in the fibrotic pancreas. The factors responsible for mediating PSC activation during chronic alcohol exposure include ethanol, its metabolite acetaldehyde, oxidant stress and cytokines (released during episodes of alcohol‐induced pancreatic necroinflammation). Most recently, the intracellular signaling mechanisms regulating ethanol‐induced PSC activation have been identified and include the mitogen‐activated protein kinase (MAPK) pathway, phosphatidylinositol‐3‐kinase (PI3K) and protein kinase C (PKC), and the transcription factor activator protein‐1 (AP‐1).
Evaluation of the role of nuclear factor‐κB signaling in liver injury using genetic animal models
Authors: Luedde Tom , Beraza Naiara , Trautwein Christian
Published: August 2006

Most chronic liver diseases are not sufficiently treatable at present and very often progress to liver fibrosis and liver cirrhosis. Several recent studies have suggested that cytokines and cytokine‐activated inflammatory signaling pathways might play an important role in the mediation of liver injury. Although pro‐inflammatory signaling pathways such as nuclear factor (NF)‐κB have evolved primarily for host defense to infections, they appear to be involved in the pathogenesis of inflammatory diseases and may mediate liver injury in response to a variety of agents and pathogens. Herein is summarized briefly some recent findings concerning the role of NF‐κB in different models of liver injury based on transgenic and knockout animal technology.
Alcohol‐induced oxidative stress and cell responses
Authors: Wu Dongmei , Zhai Qiwei , Shi Xianglin
Published: August 2006

Epidemiological and animal studies have demonstrated that alcohol abuse is directly associated with the increase of multiple organ diseases, such as liver injury, cardiovascular diseases, and neurological disorders. While the mechanisms of alcohol‐induced cell injury and disease remain to be investigated, recent studies indicate that reactive oxygen species (ROS) may play an important role. Reactive oxygen species are able to cause various cellular injuries, such as DNA damage, lipid peroxidation and protein modification. Cellular systems are protected from ROS‐induced cell injuries by an array of defenses composed of various anti‐oxidants with different functions. When the ROS present in the cellular system overpower the defense systems, they will cause oxidative stress or cell injury, leading to the development of diseases. This article reviews recent literature on alcohol‐induced ROS production, oxidative stress, signal transduction, and cellular responses. The implication of these processes in alcohol‐related diseases is also discussed.
Interactions of cytokines, S‐adenosylmethionine, and S‐adenosylhomocysteine in alcohol‐induced liver disease and immune suppression
Authors: Barve Shirish , Joshi‐Barve Swati , Song Zhenyuan , Hill Daniell , Hote Prachi , Deaciuc Ion , McClain Craig
Published: August 2006

Alcoholic liver disease (ALD) remains a leading cause of death in the USA. Defining mechanisms for liver cell death in ALD in order to develop potential new agents for therapeutic intervention is a major focus of the authors’ work. Abnormal cytokine metabolism is a major feature of ALD, and a thorough understanding of both mechanisms and interactions of cytokine overproduction and sensitization are critical to developing a possible treatment for ALD. S‐Adenosylmethionine has been used in a variety of animal studies and clinical trials and has been reported to improve biochemical parameters of liver function. Last, immunosuppression associated with chronic alcohol abuse is an important predisposing factor to opportunistic infections and cancer. It is the authors’ working hypothesis that alcohol consumption leads to chronic activation of the immune system.
Causes and consequences of mitochondrial reactive oxygen species generation in hepatitis C
Authors: Wang Ting , Weinman Steven A
Published: August 2006

Hepatitis C virus has developed mechanisms to alter the redox state of hepatocytes and this is associated with changes in mitochondrial structure and function. Chronic hepatitis C patients manifest hepatic oxidative stress and this is exacerbated by alcohol consumption and associated with fibrosis progression. Several viral proteins, including core and NS5a appear to contribute to reactive oxygen species (ROS) generation by mechanisms that involve both mitochondria and endoplasmic reticulum (ER). Hepatitis C virus (HCV) core protein localizes to both ER and mitochondria and has effects at both sites. At the mitochondria a chain of events is initiated by core binding, which consists of increased Ca2+ uptake, increased mitochondrial superoxide production, oxidation of the mitochondrial glutathione pool, inhibition of electron transport complex I activity, and sensitization of mitochondria to Ca2+‐ and ROS‐induced membrane permeability transition. These effects have been observed in isolated mitochondria, cells bearing full‐length HCV replicons, and liver mitochondria derived from HCV transgenic mice. In addition to these direct effects on mitochondria, core protein has been shown to causes a state of ER stress and an increase in the efficiency of ER to mitochondria Ca2+ transfer. The resulting oxidized redox state has a number of potential consequences for liver function. It interferes with the antiviral innate immune responses and potentiates fibrosis and carcinogenesis. Alcohol exacerbates these effects by increasing core‐induced ROS production, further oxidizing the mitochondrial glutathione pool. The resulting mitochondrial effects may contribute to liver injury and oxidative stress seen in chronic hepatitis C.