Day 1 :
Keynote Forum
Dr. Brockbank
Bioengineering, Clemson University and Tissue Testing Technologies LLC
Keynote: Cryopreservation of Complex Biomaterials
Biography:
Dr. Brockbank, CEO and Founder of Tissue Testing Technologies LLC, is a Research Professor of Bioengineering at Clemson University and Adjunct Professor of Regenerative Medicine and Cell Biology at the Medical University of South Carolina. His research interests include cell, tissue and organ cryopreservation for test systems and transplantation and manufacturing methods for cell-based bioengineered therapy products. His work has led to the establishment of two successful publicly traded low temperature technology platform companies, CryoLife, Inc., and Lifeline Scientific. He has over 500 publications and presentations at national and international conferences including more than 30 patents related to hypothermic, frozen and vitrified biomaterial preservation.
Abstract:
Effective improved tissue banking methods for natural and engineered tissues, complex vascularized allotransplants and organs are desperately needed for transplantation. Banking of living cellular tissues using current tissue banking practices employing conventional cryopreservation by freezing is not feasible due to the well documented damage caused by ice formation. An alternative ice-free cryopreservation approach is vitrification. Formation of ice is prevented by the presence of high concentrations of cryoprotectants with preservation of extracellular matrix components and optional preservation of cells. Ice-free vitrification works for a variety of natural and engineered tissues, using a formulation consisting of DMSO, formamide and propylene glycol, known as VS55, but have been unsuccessful at sample volumes over a few mLs. The major constraints for scale-up of cryopreservation by ice-free vitrification have been avoidance of ice nucleation during warming and mechanical forces generated by glasses at low temperatures. In this presentation I will focus on strategies for avoidance of ice nucleation. Our first successful strategy for large tissue samples was an 83% formulation based upon the same cryoprotectants, known as VS83. This formulation can be used to retain viable chondrocytes in large osteochondral grafts or for non-viable cardiovascular grafts with retention of extracellular matrix integrity, depending upon the way in which the formulation is added and removed before and after vitrification. Non-viable cardiovascular grafts with intact matrix have been a major research focus for the last 10 years. Both in vitro and in vivo results demonstrated significantly reduced immunogenicity in heart valves (Figure), including reduced memory T-cell proliferation and most recently modulation of TGF-β1 from latent to active form among other statistically significant effects. We have been successful in scaling up the viable preservation of large tissue samples using either nanowarming, inductive heating of iron nanoparticles, or convection warming using improved ice-free vitrification formulations incorporating low molecular weight sugars.
- Tissue Engineering, Tissue Culture & Preservation, Regenerative Medicine, Cell Therapy.
Location: WEBINAR
Session Introduction
Dr. VIKRAM PABREJA
Director. PabCyte. Gurgaon. Haryana. India
Title: Treatment of Hypothyroidism with Intrathyroidal injection of Autologous PRP and Hyaluronic Acid
Biography:
Dr. Vikram Pabreja is a physician in medical practice for last 25 years and has a rich experience in field of Stem Cell & Regenerative Medicine. He is first to demonstrate the role of stem cells in infectious diseases as dengue fever. He is Director of PabCyte, platform for advances is Stem Cell enabled therapies for degenerative, autoimmune, and infectious diseases in human population. He has published his work in Indian journals and is in process of approaching International journals as well.
Abstract:
This study was carried out to investigate the effects of intrathyroidal injection of Autologous PRP and HA under ultrasound guidance on Resident Thyrocyte Stem Cells and Thyrocyte Progenitor cells in the treatment of hypothyroidism (less T3 T4 TSH), subclinical hypothyroidism (normal T3 T4 raised TSH) and autoimmune thyroiditis. 30 patients were selected from age group of 25-70 years. All patients held oral thyroxine tablets prior to intrathyroidal injection of autologous PRP and HA. 20 ml Peripheral venous blood was withdrawn after pharmacologic intetvention with iv Omega 3 fatty acids 10% 5gm and iv ascorbic acid without preservative 5gm. Further it was centrifuged at 3300 rpm for 7 minutes to yield 6ml of PRP. This PRP was mixed with 1ml of Hyaluronic Acid, 22 mg/ml, non-avian & non-allergenic. PRP was tested for microbial contamination before injection. 3ml of PRP and 1ml of HA was injected in each lobe of thyroid gland under ultrasound guidance. The patients were followed up for a period of 1 year with 1 month, 3 months and 6 months and 1 year interval. Thyroid function Test, free T3, free TSH anti TPO, anti TG antibody tests were assessed along with ultrasound of thyroid gland. Of the 30 patients 12 were Males and 18 females. Females were categorized into 3 adoloscent (age 14-19 years)/TSH 7-15 micro IU/ml 10 (age 21-40 years)/TSH 10-30 micro IU/ml, 5 (Age 41-60 years)/TSH 8-16 micro IU/ml. A total of 3 injections were given at interval of 15 days. RESULTS 27 patients responded well to the treatment and improved after 1 month of injection. No adverse events were found during 1 year follow up with significant decrease in serum TSH, anti TPO and anti TG antibody values. T3 T4 increased in primary hypothyroidism situations. Patients reported less lethargy less musculoskeletal pains, better concertration, improved mood improvement in dry skin and hair loss and no weight gain from baseline. CONCLUSION It is evident that thyroid niche stem cells / Progenitor cells on stimulation have regenerative potential. Either new thyrocytes come up in the gland or earlier non functional or hypofunctioning thyrocytes get repaired and resume normal function. Thus intrathyroidal injection of autologous PRP and HA under ultrasound guidance can be a safe, better and effective therapy that can permanently rescue thyroid function. PRP and HA together act as a bioscaffold.
Dr. Nikolai Sopko
Nikolai Sopko, MD, PhD, PolarityTE, USA
Title: Complex cutaneous wounds treated with autologous homologous skin construct yields neo-generation of functional full-thickness skin in preclinical and clinical models
Biography:
Dr. Nikolai Sopko obtained his MD with AOA honors from Case Western Reserve University and PhD in stem cell biology at the Cleveland Clinic as a graduate of the NIH Medical Scientist Training Program. He completed his surgical training at the Johns Hopkins James Buchanan Brady Urologic Institute as a Physician Scientist Scholar where he focused on regenerative reconstruction, tissue engineering, gene therapy, cell/molecular biology, transplant modulation and tumor biology. He has over 60 published peer-reviewed articles and book chapters and is a speaker on regenerative medicine. Dr. Sopko is currently the Chief Scientific Officer and Vice President of Research and Development of PolarityTE where he applies his passion for patient care and basic science research to develop novel regenerative therapeutics for clinically challenging diseases.
Abstract:
Critical cutaneous wounds affect thousands of individuals each year. Skin requires regeneration of all components to function normally after such traumas. Current established standards of care, such as split-thickness skin grafts (STSG), do not allow for full regeneration or function of skin and often result in incomplete healing, scarring, and contraction. SkinTE™ is a novel autologous homologous skin construct (AHSC) that has the ability to regenerate full-thickness skin with associated appendages, including sweat glands and hair follicles. AHSC treats a full spectrum of conditions of the skin in a manner that promotes and utilizes self-regeneration early in the healing process. AHSC was evaluated in both adult and pediatric patients for its ability to close complex wounds in multiple clinical settings. RAMAN spectroscopy and macro and microscopic imaging were used to analyze neo-generation of dermal appendages, and AHSC shows promise for the regeneration of full-thickness hair-bearing skin. Preclinical data shows regeneration of epidermis, dermis, and dermal appendages with decreased contraction and scarring. Clinical applications have shown complete healing in complex cutaneous wounds, including pigment development and appendage regeneration. Improved function, sensation, contour, and superior cosmesis as compared to STSGs were also noted. We compare clinical outcomes of AHSC to current standard of care therapies for critical skin wounds and summarize the functional outcomes from a large case series following the application of AHSC in early clinical settings for a multitude of complex wounds. We confirmed the generation of dermal appendages with multiple imaging and observational platforms and evaluated the molecular profile of AHSC compared to native skin. Lastly, we evaluated macro-properties of AHSC including collagen content, fibril organization, and appendage morphology compared to native skin. We concluded that one application of AHSC has the capacity to facilitate the regeneration of full-thickness, fully functional skin and reduces the surgical and healthcare burden for acute and chronic cutaneous wounds, acute burns, burn reconstruction, scar revision, surgical reconstruction, and the replacement of skin grafts.
Syed Muntazir Mehdi
University of Agriculture, Faisalabad, Pakistan.
Title: Suppression of begomoviruses using CRISPR/Cas9 with multiple gRNAs
Biography:
Syed Muntazir Mehdi has completed his Bachelor in Bioinformatics and Biotechnology at the age of 22 years from GC University, Faisalabad ,Pakistan and Master studies from University of Agriculture, Faisalabad,Pakistan.
Abstract:
Site-specific genome editing through CRISPR/Cas has revolutionized the field of plant genome engineering. CRISPR/Cas is an effective, efficient and economical method of targeted genome engineering in almost every field of biology and agriculture. In addition, targeting multiple genes simultaneously has made Cas9 a method of choice for functional genomics and engineering resistance in crops against biotic and abiotic stresses. We designed multiple gRNAs to target simultaneously three genes of cotton leaf curl virus for viral inference in plant. In transient assay, replication of the virus was halted by Cas9 having multiple gRNAs leading to low virus titer and attenuated viral symptoms. Cotton transformation was optimized with DsRED and stable transformation of Cas9 along with multiple gRNAs in cotton is in progress. Validation of multiple gRNAs for virus suppression will open up new corridors for using Cas9 for suppression of viral diseases in crop plants.
Mahdieh Heydarigoojani
university of Vienna,Austria
Title: On the mechanism of angiogenesis induced by thrombin receptor activating peptides(TRAP-9)
Biography:
Mahdieh Heydarigoojani studied Biomedical engineerinmg and intrested in improvement of angiogenesis and vascularization in tissue engineering field. She graduated in 2014 and recived her bachelor in International Conference on Fibromyalgia and Chronic Pain (June 15-16, 2016 Philadelphia, USA) J Pain Relief ISSN: 2167-0846 JPAR an open access journa Fibromyalgia 2016 June 15-16, 2016 Volume 5, Issue 3(Suppl) Page 40 Biomedical engineering. She intended to improve her knowledge and accepted at technical university of Vienna, studied Biomedical engineering in master. Due to personal passion for tissue engineering, she decided to work in this field and started her master thesis in tissue engineering with focus on improvement of vascularozation at LBI institute. She would like to continue her carrier in this way.
Abstract:
Angiogenesis and vascularization is critical issue in tissue engineering. Thrombin, a serin protease, plays an important role in inflamation and injury. It is present in site of inflammation and activates platelet and different cellular response including angiogenesis. It interact with protease activated receptors (PAR) on endothelial cell surface and cleaves terminal portion of receptor which new N-terminal autostimulates the receptor and cause transmembrane signaling. Synthetic peptide containing agonist motif such as SFLLRNCCC called thrombin receptor activating preptides (TRAP-9) for human PAR-1, capable of causing full receptor activation and mimic many effects of thrombin. In current study, we investigate the maximum concentartion of TRAP-9 which is capbale of human umbilical vain endothelial cell (HUVEC) proliferation. Diffreent HUVEC cell marker´s expression after stimulation with TRAP-9 and thrombin were determined. Also effect of TRAP-9 on vascular network formation in HUVEC/ASC co-culture was assessed. Method In this study, thrombin receptor activating peptide(TRAP-9) with motif SFLLRNCCC(Ser,Phe,Leu,Leu,Arg,Asn,3*Cys) was synthetized.poweder was diluted with PBS and and freezed at -20oc . confluent HUVEC was treated with different TRAP-9 concentartion and proliferation after 24h wa checked with ELISA.Different HUVEC cell marker was determined using flow cytometry after 24 stimulation with TRAP-9 and thrombin.Finally effect of TRAP-9 on vascular tube formation in GFP-HUVEC/ASC coculture was assessed and networks were quantified using AngioSys. Result Proliferation of HUVEC enhanced with TRAP-9 which 500µM showed higher proliferation compare to lower concentartions.HUVECs markers expression level is in similar amount as thrombin. Number of junctions,tubuls and total tubul length improved when cells terated with TRAP-9. Conclusion Data showed that TRAP-9 is capable HUVEC proliferation without toxic effect even in high concentartion.Level of cell markers expression is comparable with thrombin. Incorporation of TRAP-9 in fibrin clot co-culture indicates that this motif improved networks formation and would be promissing agonist for thrombin for angiogenesis. Key words: HUVEC/ASC co-culture,TRAP-9,PAR-1,angiogenesis,thrombin