Day 2 :
CEO & Founder - Biotechnology and Regenera-tive Medicine at RegenerAge International
Keynote: RegenerAge System: Therapeutic effects of combinatorial biologics (Bioquantine®) and Spinal cord stimulation system on a patient with Spinal Cord section
Time : 10:00 - 10:45
CEO & Founder - Biotechnology and Regenera-tive Medicine at RegenerAge International.VP of International Clinical Development for Bioquark, Inc. Chief Clinical Officer at ReAnima™ Advanced Biosciences. Westhill University School of Medicine. Mexico.Advance Fellow by the American Board of An-tiAging and Regenerative Medicine (A4M).Visiting scholar at University of North Carolina at Chapel Hill (Dermatology).Fellow in Stem Cell Medicine by the American Academy of Anti-Aging Medicine and Universi-ty of South Florida.
As it has been previously demonstrated that co-electroporation of Xenopus laevis frog oocytes with normal cells and cancerous cell lines in-duces the expression of pluripotency markers, and in experimental murine model studies that Bioquantine® extract (purified from intra- and extra-oocyte liquid phases of electroporated oocytes) showed potential as a treatment for a wide range of conditions as Squint, Spinal Cord Injury (SCI) and Cerebral Palsy among others. The current study observed beneficial changes with Bioquantine® administration in a patient with a severe SCI. Pluripotent stem cells have therapeutic and regenerative potential in clinical situations CNS disorders even cancer.2-3-7 One method of reprogramming somatic cells into pluripotent stem cells is to expose them to ex-tracts prepared from Xenopus laevis oocytes1 We showed previously that coelectroporation of Xenopus laevis frog oocytes; with normal cells and cancerous cells lines, induces expression of markers of pluripotency.4 We also observed ther-apeutic effects of treatment with a purified ex-tract (Bioquantine) of intra- and extra-oocyte liquid phases derived from electroporated X. laevis oocytes, on experimentally induced pathologies including murine models of melanoma, traumatic brain injury, and experi-mental skin wrinkling induced by squalene-monohydroperoxide (Paylian et al, 2016). The positive human findings for Spinal Cord Injury, and Cerebral Palsy with the results from previ-ous animal studies with experimental models of traumatic brain injury, respectively (Paylian et al, 2016). Because of ethical reasons, legal re-strictions, and a limited numbers of patients, we were able to treat only a very small number of patients. These results indicate that Bioquan-tine® may be safe and well tolerated for use in humans, and deserves further study in a range of degenerative disorders. We propose that the mechanism of action of Bioquantine® in these various diseases derives from its unique phar-macology and combinatorial reprogramming properties. In conclusion, these preliminary find-ings suggest that Bioquantine is safe and well tolerated on patients with Cerebral Palsy and-Spinal Cord Injury, among others. In addition to the regenerative therapy and due to the patient condition, we decided to include the Restore-Sensor SureScan5-6 . Based on the of electrical stimulation for rehabilitation and regeneration after spinal cord injury published by Hamid and MacEwan 8-9 , we designed an improved deliv-ery method for the in situ application of MSCs and Bioquantine® in combination with the RestoreSensor® SureScan® Conclusions: To the present day the patient who suffered a total sec-tion of spinal cord at T12-L1 shows an im-provement in sensitivity, strength in striated muscle and smooth muscle connection, 9 months after the first therapy of cell regeneration and 1 month after the placement of RestoreSensor® at the level of the lesion, the patient with a com-plete medullary section shows an evident im-provement on his therapy of physical rehabilita-tion in standing for the first time and showing a progressively important functionality.
- Tissue Repair and Regeneration | Tissue Engineering | Epigenome and Epigenetic Analysis | Biomaterials and Bioengineering
Joel I Osorio
Elizabeth Pavez Loriè has her expertise in skin biology, dermatology and tissue culturing and a passion in using in vitro modeling systems to provide answers to dermatological conditions and disease, linking biology to medicine and vice versa. Her use of models started when she used in vitro models to understand the effects of retinoic acid blocking agents as well as the function of specific cytochromes in the human epidermis and additionally the first steps to use a disease model to study drug related outcomes (6, 7). She then focused on long-term skin equivalents and non-melanoma skin cancer and aging, specifically the impact of the sun on human skin, working in close collaboration with Prof. Petra Boukamp. Her goal is to provide robust and biologically accurate models to investigate the homeostatic state of the skin as well as to understand and provide the clinic with personalized testing long-term models. To do this she would like to continue developing the skin models to meet the needs of the biological/medical questions in hand.
An important part for the construction of skin equivalents is the living dermal compartment, which includes fibroblasts these are mainly seeded and supported by different structures (DED, collagen, Matrigel, scaffold etc). Another type of dermal equivalent, developed by Ahlfors and Billiar (2) and later adapted by Berning et al (1) among others is the cell derived matrix (cdm) model. It has no “artificial” support and is also well suited for tumor invasion studies. Here the tumorigenic cells’ behavior can be examined and their specific invasion pattern can be followed. Most skin models have a short life span and are mostly suited and used for studying acute effects, but many of the environmental agents or dermatological conditions that face the skin will not only have an immediate effect. To be able to mimic and follow conditions over a longer period of time we base our studies on stable skin equivalents that allow the generation and regeneration of epithelial tissue for up to 24 weeks (1-5), allowing us to understand the long term effects of any given alteration. The sun is one of the key environmental factors affecting
the skin with our cdm model we are able to follow the effects of chronic UV exposure for several months. We have also been able to mimic “old” skin and follow the long-term effects of experimental sunlight on this aging model. Additionally this non-immunogenic model, suits perfectly well to examine the effects of drugs such as Cyclosporin A, mimicking the skin’s condition in organ transplant recipients over a longer period of time. The use diversity makes these models great members of the skin equivalent family and hopefully they continue to contribute in studies and in the future give us an insight into the biological dimension of time.
Wake Forest Institute for Regenerative Medicine, USA
Ronald A. Nelson, Jr. earned his Ph.D in Chemistry from Wake Forest University where he designed, synthesized and evaluated the effectiveness of phosphatidylinositol 3 kinase inhibitor prodrugs in treating androgen - independent prostate cancer. Serving in over 11 different leadership roles during his graduate career, Ronald's commitment to excellence led to 4 national and 15 local awards. Passionate about research, medicine, technology, his move to the Wake Forest Institute for Regenerative Medicine was a natural fit. As a postdoctoral research fellow at WFIRM, Ronald has been the research lead for several animal surgical, microsurgical, and treatment procedures. Ronald is also the research manager for the Regen Med Development Organization (ReMDO) where he utilizes his background in analytical chemistry, cancer biology, organic synthesis, and engineering to develop universal cell culture media that is chemically-defined and xeno-free. Ronald plans to enhance this media to recapitulate the regenerative potential of cells observed in embryogenesis.
Background: Animal-derived biological extracts, including animal serum, are commonly used in tissue culture medium to provide bioregulatory factors that support the maintenance of cell viability and promote cell proliferation. These biological extracts are not chemically defined, are inconsistent from lot to lot, and carry the risk of disease transmission. As such, these media supplements are not optimal materials for use in clinical manufacturing processes. Our group has developed a chemically defined media based on the known constituents within the well characterized biological extract human platelet lysate. This media has been shown to provide exceptional support for most cell types derived from the mesodermal embryonic germ layer.
Methods: Human sourced or recombinant versions of the major bioregulatory factors present in human platelet lysate were added to a modified DMEM-F12 minimal medium. The factors added included platelet-derived growth factor, transforming growth factor-β, insulin-like growth factor, vascular endothelial growth factor, fibroblast growth factor, hepatocyte growth factor, epidermal growth factor, and several other factors. Growth curves for an extensive panel of commercially available human primary cells were generated using an IncuCyte S3 Live Cell Imager. Preservation of cell phenotype was confirmed by immunofluorescent determination of cell type specific functional proteins.
Results: Proliferation rates for most mesoderm derived cell types in the chemically defined medium were equivalent or superior to proliferation rates measured in the cell supplier’s recommended, chemically undefined medium. Preservation of functional biomarker expression indicated that cell phenotype was maintained across multiple cell passages for each cell type. However, two endothelial cell types, human umbilical vascular endothelial cells and human dermal microvascular cells, did not proliferate in the chemically defined medium.
Conclusion: A chemically defined cell culture medium based on the known constituents of the biological extract, human platelet lysate was formulated using human sourced and recombinant protein bioregulatory factors. This media formulation was shown to support proliferation and preservation of phenotype for most cell types of mesodermal origin. Because this medium is chemically defined and xeno-free, it represents an optimal reagent for use in clinical manufacturing processes.
PhD student in the Helmholtz Group for Cell Biology
Title: The seminiferous tubules of mammalian testes: A different epithelium encased by a bandage structure of smooth muscle cell monolayers
Time : 12:05-12:35
Lisa M. Domke is a PhD student in the Helmholtz Group for Cell Biology of Professor Werner W. Franke (German Cancer Research Center, DKFZ, Heidelberg, Germany). Before she has prepared her master thesis with one of the pioneers in cancer research, Prof. Dr. Robert A. Weinberg, at the Massachusetts Institute of Technology (MIT) in Cambridge, USA. The nature of her degree in Biotechnology has allowed her to learn various analytical as well as light and electron microscopical techniques and to work in different fields of life sciences. At present she is finishing her thesis at the DKFZ with Prof. Dr. Werner W. Franke on the molecular and ultrastructural characterization of cell-cell junctions and cytoskeletons in the tubuli seminiferi and their encasing contractile peritubular wall smooth muscle system of diverse mammalian species to improve current methods of molecular diagnostics.
Mature seminiferous tubules (STs) of mammalian testes comprise the Sertoli cells and germ cells and are tightly surrounded by a special peritubular cell wall. Using biochemical, immunocytochemical and electron microscopical methods, we have determined that STs differ from all other epithelia by the absence of cytokeratin intermediate filaments (IFs) but are rich in vimentin IFs, do not contain major epithelial marker structures and molecules such as desmosomes or E-cadherin-based adherens junctions (AJs) but contain exclusively N-cadherin-based AJs. In Sertoli cells, we have found two new junction structures:(i)N-cadherin-based areae adhaerentes which often represent even very large areas connecting Sertoli cells with each other or with germ cells. (ii) Special AJs arranged in closely and regularly spaced rows of tight junction-like structures and associated with 5-8 nm wide cytoplasm-to-cytoplasm channels(“cribelliform junctions”).
The seminiferous tubule cells are attached to the peritubular wall by a well-developed basal lamina but lack hemidesmosomes and hemidesmosomal marker molecules. The peritubular wall is a stack system of layers of extracellular matrix (ECM) structures alternating with monolayers of very flat “lamellar smooth muscle cells” (LSMCs). These LSMCs represent differentiated smooth muscle cells (SMCs; positive for smooth muscle α-actin, the corresponding myosin light and heavy chains, α-actinin, tropomyosin, smoothelin, desmin, vimentin, filamin, talin, dystrophin, caldesmon, calponin and protein SM22α). The cells are laterally connected – often in overlapping protrusions – by AJs containing cadherin-11 as the predominant cadherin, and also P-cadherin and rarely N-cadherin, anchored in cytoplasmic plaques containing β-catenin, proteins p120 and p0071, plakoglobin and protein myozap. LSMCs also contain typical SMC structures such as “dense bodies”, plasma membrane-associated “focal adhesions” and caveolae. Thus, we conclude that these LSMCs represent a specific SMC type and not just “myoid cells” or myofibroblasts as stated in the literature.