In addition to its normal function, the adenine nucleotide translocase (ANT) forms the inner membrane channel of the mitochondrial permeability transition pore (MPTP). Binding of cyclophilin-D (CyP-D) to its matrix surface (probably on Pro 61 on loop 1) facilitates a calcium-triggered conformational change converting it from a specific transporter to a non-specific pore. The voltage dependent anion channel (VDAC) binds to the outer face of the ANT, at contact sites between the inner and outer membranes, and together VDAC, ANT and CyP-D probably represent the minimum MPTP configuration. The evidence for this is critically reviewed as is the structure and molecular mechanism of the carrier in its normal physiological mode. This provides helpful insights into MPTP regulation by adenine nucleotides, membrane potential and ANT ligands such as carboxyatractyloside and bongkrekic acid. Oxidative stress activates the MPTP by glutathione-mediated cross-linking of Cys 159 and Cys 256 on matrix-facing loops of the ANT that inhibits ADP binding and enhances CyP-D binding. Molecular modeling of the loop containing the ADP binding site suggests an arrangement of aspartate and glutamate residues that may provide a calcium binding site. There are other proteins that may bind to the ANT, modulating MPTP opening and hence cell death. These included members of the Bax/Bcl-2 family (both oncoproteins and tumor suppressors) and viral proteins. Vpr from HIV-1 can bind to ANT and convert it into a pro-apoptotic pore, whereas vMIA from cytomegalovirus interacts to inhibit opening. Thus the ANT may provide a molecular link between physiopathological mechanisms of infection and the regulation of MPTP function and so represents a potential therapeutic target.

The adenine nucleotide translocase: A central component of the mitochondrial permeability transition pore and key player in cell death : Mitochondria as a target for apoptosis modulation

Pharmacognostic and pharmacological profile of Humulus Lupulus L.

In seven strains of cultured normal human osteoblast-like cells, a mean of 1615 molecules of tritium-labeled 17 beta-estradiol per cell nucleus could be bound to specific nuclear sites. The nuclear binding of the labeled steroid was temperature-dependent, steroid-specific, saturable, and cell type-specific. These are characteristics of biologically active estrogen receptors. Pretreatment with 10 nanomolar estradiol in vitro increased the specific nuclear binding of progesterone in four of six cell strains, indicating an induction of functional progesterone receptors. RNA blot analysis demonstrated the presence of messenger RNA for the human estrogen receptor. The data suggest that estrogen acts directly on human bone cells through a classical estrogen receptor-mediated mechanism.

Evidence of estrogen receptors in normal human osteoblast-like cells

mTOR complex 2 (mTORC2) contains the mammalian target of rapamycin (mTOR) kinase and the Rictor regulatory protein and phosphorylates Akt. Whether this function of mTORC2 is critical for cancer progression is unknown. Here, we show that transformed human prostate epithelial cells lacking PTEN require mTORC2 to form tumors when injected into nude mice. Furthermore, we find that Rictor is a haploinsufficient gene and that deleting one copy protects Pten heterozygous mice from prostate cancer. Finally, we show that the development of prostate cancer caused by Pten deletion specifically in prostate epithelium requires mTORC2, but that for normal prostate epithelial cells, mTORC2 activity is nonessential. The selective requirement for mTORC2 in tumor development suggests that mTORC2 inhibitors may be of substantial clinical utility.

mTOR Complex 2 Is Required for the Development of Prostate Cancer Induced by Pten Loss in Mice

Ischemia/reperfusion injury is an unavoidable relevant consequence after kidney transplantation and influences short term as well as long-term graft outcome. Clinically ischemia/reperfusion injury is associated with delayed graft function, graft rejection, chronic rejection and chronic graft dysfunction. Ischemia/reperfusion affects many regulatory systems at the cellular level as well as in the renal tissue that result in a distinct inflammatory reaction of the kidney graft. Underlying factors of ischemia reperfusion include energy metabolism, cellular changes of the mitochondria and cellular membranes, initiation of different forms of cell death-like apoptosis and necrosis together with a recently discovered mixed form termed necroptosis. Chemokines and cytokines together with other factors promote the inflammatory response leading to activation of the innate immune system as well as the adaptive immune system. If the inflammatory reaction continues within the graft tissue, a progressive interstitial fibrosis develops that impacts long-term graft outcome. It is of particular importance in kidney transplantation to understand the underlying mechanisms and effects of ischemia/reperfusion on the graft as this knowledge also opens strategies to prevent or treat ischemia/reperfusion injury after transplantation in order to improve graft outcome.

Update on ischemia-reperfusion injury in kidney transplantation: Pathogenesis and treatment

New insight in the neuropharmacological activity of Humulus lupulus L.

Influence of Eurycoma longifolia on the copulatory activity of sexually sluggish and impotent male rats.

The main aim of this study is to evaluate potential human stem cells, such as dental pulp stem cells and amniotic fluid stem cells, combined with collagen scaffold to reconstruct critical-size cranial bone defects in an animal model. We performed two symmetric full-thickness cranial defects on each parietal region of rats and we replenished them with collagen scaffolds with or without stem cells already seeded into and addressed towards osteogenic lineage in vitro. After 4 and 8 weeks, cranial tissue samples were taken for histological and immunofluorescence analysis. We observed a new bone formation in all of the samples but the most relevant differences in defect correction were shown by stem cell–collagen samples 4 weeks after implant, suggesting a faster regeneration ability of the combined constructs. The presence of human cells in the newly formed bone was confirmed by confocal analysis with an antibody directed to a human mitochondrial protein. Furthermore, human cells were found to be an essential part of new vessel formation in the scaffold. These data confirmed the strong potential of bioengineered constructs of stem cell–collagen scaffold for correcting large cranial defects in an animal model and highlighting the role of stem cells in neovascularization during skeletal defect reconstruction.

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Human amniotic fluid-derived and dental pulp-derived stem cells seeded into collagen scaffold repair critical-size bone defects promoting vascularization

The cartilage tissue engineering associated with stem cell-related therapies is becoming very interesting since adult articular cartilage has limited intrinsic capacity for regeneration upon injury. Amniotic fluid stem cells (AFSC) have been shown to produce exosomes with growth factors and immunomodulating molecules that could stop tissue degradation and induce cartilage repair. Based on this state of the art, the main aim of this study was to explore the efficacy of the secreted exosomes, compared to their AFSC source, in MIA-induced animal model of osteoarthritis mimicking a chronic and degenerative process, where inflammation is also involved and lead to irreversible joint damage. Exosomes, obtained by the use of a commercial kit, prior to the injection in animal knee joints, were characterized for the presence of typical markers and HGF, TGFβ, and IDO. Then, analyses were performed by histology, immunohistochemistry, and behavioral scoring up to 3 weeks after the treatment. Exosome-treated defects showed enhanced pain tolerance level and improved histological scores than the AFSC-treated defects. Indeed by 3 weeks, TGFβ-rich exosome samples induced an almost complete restoration of cartilage with good surface regularity and with the characteristic of hyaline cartilage. Moreover, cells positive for resolving macrophage marker were more easily detectable into exosome-treated joints. Therefore, a modulating role for exosomes on macrophage polarization is conceivable, as demonstrated also by experiments performed on THP1 macrophages. In conclusion, this study demonstrates for the first time the efficacy of human AFSC exosomes in counteract cartilage damage, showing a positive correlation with their TGFβ content.

Manuela Zavatti

Papers

Pharmacognostic and pharmacological profile of Humulus Lupulus L.

New insight in the neuropharmacological activity of Humulus lupulus L.

Influence of Eurycoma longifolia on the copulatory activity of sexually sluggish and impotent male rats.

Human amniotic fluid-derived and dental pulp-derived stem cells seeded into collagen scaffold repair critical-size bone defects promoting vascularization

Comparison of the therapeutic effect of amniotic fluid stem cells and their exosomes on monoiodoacetate-induced animal model of osteoarthritis.