MicroRNA target homeobox messenger RNA in HIV induced hematopoietic inhibition.

Cytopenias are a common occurrence due to abnormal hematopoiesis persistent in patients suffering from and advancing with HIV/AIDS. In order to develop efficacious therapies against cytopenias, it is necessary to understand the mechanisms by which HIV infection affects the differentiation of hematopoietic stem-progenitor cells (HSPCs), causing hematopoietic inhibition, that leads to hematological disorders. Currently, only the antiretrovirals that are being used to treat HIV infection and indirectly lower the levels of virus replication also co-attenuate cytopenias. The evidence available suggests that this indirect efficacy may not prevail for the lifetime of the infected patients, and the acquired immunodeficiency can overtake the beneficial consequences of decreased virus replication. As cited in this article, we and our colleagues are the first to make a foray into the involvement of microRNAs and their use as potential interventional treatments for the cytopenias that occur with HIV/AIDS. Herein, we progressed further in the direction of the mechanisms of the involvement of homeobox gene regulation to cause cytopenias. We had previously shown that HIV-1 inhibits multi-lineage hematopoiesis of the CD34+ cells using SCID-hu Thy/Liv animals in vivo. Furthermore, we demonstrated that the virus-induced hematopoietic inhibition occurred despite the CD34+ cells being resistant to HIV-1 infection. We set out to search for the specific host factors secreted by CD4+ T-cells that likely participate in the inhibition of hematopoiesis of the HIV infection-resistant CD34+ cells. More recently, we reported the identification of virus-infected CD4+ thymocyte-secreted miRNA-15a and miRNA-24 and that their differential expression following HIV infection causes the indirect inhibition of hematopoiesis. We then hypothesized that the observed miRNA differential expression in the virus-infected T-cells causes the abnormal regulation of homeobox (HOX) gene-encoded transcriptomes in the CD34+ cells, affecting specific MAPK signaling and CD34+ cell fate, thereby disrupting normal hematopoiesis. We present that in HIV infection, miRNA-mediated post-transcriptional dysregulation of HOXB3 mRNA inhibits multi-lineage hematopoiesis, which translates into hematological disorders in virus-infected patients with HIV/AIDS. These observations portend specific microRNA candidates for potential efficacy against the virus-induced cytopenias that are otherwise not treatable by the existing HAART/ART regimens, which are primarily designed and applicable for the attenuation of virus replication.

Contrasting mechanistic susceptibilities of hematopoietic and endothelial stem-progenitor cells in respective pathogeneses of HIV-1 and SARS-CoV-2 infections.

The multitude of cellular types can be expected to behave differently when receiving invading pathogens such as mammalian viruses. The nature-dictated causes for such intrinsic cellular diversity become the criteria for the emergence of specific virus-receptor interactions on that particular host cellular surface, in order to accommodate contact with various other living entities whether desirable to the host or not. At present, we are presented with an example of two contrasting behaviours wherein the well-known HIV-1 and the more recently emergent SARS-CoV-2 cause adverse consequences to the differentiation and functions of progenitor stem cells. These include the two different downstream multipotent CD34+ hematopoietic (HSPC) and CD133+ endothelial (ESPC) stem-progenitor cells of their common pluripotent hemangioblast precursors. The two viruses target the respective endothelial and hematopoietic stem-progenitor cells to thrive upon the relevant host cellular surrounded stromal microenvironments by adopting reciprocally-driven mechanistic routes, which incidentally cause pathogenesis either directly of ESPC (SARS-CoV-2), or indirectly of HSPC (HIV-1). HIV-1 utilizes the CD4+ T-lymphocyte receptor thereby advancing pathogenesis indirectly to the CD34+ HSPC. SARS-CoV-2 directly targets the CD133+ ESPC via ACE2 receptor causing cytokine storms of the CD4+ T-lymphocytes. In this manner, these two viruses cause and extend their damage to the other cellular sub/types coexisting in the host cellular microenvironments. The infected individuals require clinical interventions that are efficacious to prevent cellular dysfunction and ultimate cell depletion or death. We infer from these viruses mediated pathogeneses mechanisms a potential common origin of microRNA molecular therapies to address cellular dysfunctions and prevent cell loss.

HIV-1 inhibits haematopoiesis via microRNA secreted by virus-infected CD4+ T cells.

INTRODUCTION: HIV-1-infected patients develop haematological disorders such as cytopenias. One possible explanation is the inhibition of haematopoiesis at the level of differentiation of CD34+ haematopoietic progenitor stem cells. Based on our previous studies, we hypothesised that there may be viral encoded, or host cellular factors which participate in the process of inhibition of haematopoiesis. MATERIALS AND METHODS: Virus-depleted media from infected CD4+ T cells was prepared by filtration and added to CD34+ cell differentiation semisolid medium. We have also used the virus-depleted media to isolate host/viral factors including miRNA. Isolated miRNAs were screened for their haematopoietic inhibitory function using the miRNA mining approach. RESULTS: Addition of virus-depleted media caused a 40% inhibition of differentiation of CD34+ cells into myeloid and erythroid colony formation. Real-time RT-PCR showed miR-15a and miR-24 from both pIndie-C1 and pNL4.3 HIV-1-infected cells showed a significant differential expression when compared to control media. CONCLUSION: In this study, we have identified two miRNAs, miR-15a and miR-24 secreted from purified HIV-1-infected CD4+ T cells that inhibited CD34+ haematopoietic progenitor stem cell differentiation into myeloid and erythroid colonies in vitro.

Evaluation of Bone Marrow Processing Protocol for Therapeutic Applications via Culture and Characterization of Mesenchymal Stem Cells.

INTRODUCTION: Human mesenchymal stem cells from bone marrow (hMSCs) have broad therapeutic potential. These cells can be are readily isolated from bone marrow by their property to adhere to tissue culture treated culture wares. However, the proliferation rates and other properties of the cells gradually change during expansion. This study aims to validate the protocol of isolation and differentiation of hMSCs from bone marrow for therapeutic applications. METHODS: Sixty ml of bone marrow was extracted from 5 patients and MSCs were isolated. These were characterized by Flow Cytometry, CFU assay and were differentiated into bone, fat cells and neurocytes. RESULTS: The cells were having healthy morphology. These were positive for the markers CD105, CD90 and CD73 and negative for CD45, CD34 and HLA-DR. The cells could differentiate into fat, bone and neural cells. CONCLUSION: MSCs from the bone marrow were isolated and differentiated. These cells were morphologically healthy and passed CFU assay. The cells exhibited differentiation potential into bone, fat and neural tissue. These cells can be used in therapeutic applications.

A Short Study Report on Bone Marrow Aspirate Concentrate Cell Therapy in Ten South Asian Indian Patients with Autism.

Autism is a neural disorder presenting in the early developmental period, usually in the first 2 years of life. It is characterized by persistent deficits in social communication and social interaction, restricted and repetitive patterns of behavior, interests, or activities, and causes clinically significant impairment in social, occupational, or other important areas of functioning.Cellular therapy is an advanced approach to treat disorders where current therapies do not offer a cure or efficient symptom relief. Herein we present the data of ten autistic patients who were enrolled in this study utilizing stem cells. All patients were scored on ISAA and WeeFIM scales before and at 3, 6, 12 or 24 months following the injection of stem cells. The ISAA scores of the patients improved with the injection, and no adverse effects were noted. We report promising results in this small pilot study using autologous Bone Marrow Aspirate Concentrate (BMAC) injected intrathecally into these ten enrolled autistic patients.

Therapy with Bone Marrow-Derived Autologous Adult Stem Cells in Quadriparesis due to Motor Neuron Disease.

OBJECTIVE: To report the safety and therapeutic effectiveness of application of concentrated bone marrow aspirate in three bedridden patients with weakness in both legs, and monitor potential improvement in neurological outcomes. DESIGN: Case report. Intervention: Five infusions of 3x108 mononuclear cells were administrated with 12 week intervals. Bone marrow (240ML) were obtained from the posterior superior iliac spine and Bone marrow mononuclear cells were enriched by standard manual close method under aseptic condition. RESULTS: During the follow-up study of one year after stem cell implantation, the conditions of all three patients were improved and were confirmed by physical assessment, muscle charting and Electromyography (EMG). One year after stem cell implantation patients who were bedridden before treatment could sit without support and walk with support up to 200 feet at a stretch. CONCLUSION: The local application of a cocktail of regenerative cell population found in an MNC fraction of bone marrow was safe and effective in improving quality of life and muscle strength in ALS patients. This case opens the need for further investigations on Autogenic stem cell transplant therapies for MND disease.

Administration of Autologous Bone Marrow-Derived Stem Cells for Treatment of Cerebral Palsy Patients: A Proof of Concept.

BACKGROUND: Stem cell therapy is a promising treatment for cerebral palsy, which refers to a category of brain diseases that are associated with chronic motor disability in children. Autologous bone marrow stem cells may be a better cell source and have been studied for the treatment of cerebral palsy because of their functions in tissue repair and the regulation of immunological processes. METHODS: To assess autologous marrow stem cells as a novel treatment for patients with moderate-to-severe cerebral palsy, a total of 10 cerebral palsy patients were enrolled in this clinical study with 24 months follow-up. A total of 10 cerebral palsy patients received autologous bone marrow cells transplantation (4.5 × 108 mononuclear cells; 90% viability) into the subarachnoid cavity and rehabilitation. RESULTS: We recorded the gross motor function measurement scores, manual ability function measurement score, and adverse events up to 24 months post-treatment. The gross motor function measurement scores were significantly higher at month 6 post-treatment compared with the baseline scores and were stable up to 24 months follow-up. The increase in manual ability and communication function measurement scores at 6 months were not significant when compared to the baseline score. All the 10 patients survived and none of the patients experienced any serious adverse events or complications. CONCLUSION: Our results indicated that bone marrow derived MNCs are safe and effective for the treatment of motor deficits related to cerebral palsy. Further randomized clinical trials are necessary to establish the efficacy of this procedure.

Platelet-Rich Plasma Growth Factor Concentrated Spray (Keratogrow®) as a Potential Treatment for Androgenic Alopecia.

The objective is to investigate the safety and clinical efficacy of Autologous Platelet Rich Plasma Concentrated Spray (Keratogrow®), for hair loss. Autologous -PRP spray, prepared from a small volume of blood, was applied on the selected patients' scalps at least twice daily. Three months treatments were given for each patient. The effectiveness of the medication was measured by changes in hair regrowth after 3 months determined by physical exam and digital photography. At the end of the 3 cycles of treatment, the patients presented clinical improvement in the mean number of hairs, with a mean increase of hairs in the target area, and a mean increase in total hair density compared with baseline values.

Autologous Bone Marrow-Derived Stem Cells in Spinal Cord Injury.

INTRODUCTION: Spinal cord injury is a traumatic neurological condition which makes the patient disable. Its management still remains challenging but advancements in the regenerative medicine have changed the approach of treating this serious debilitating condition of the central nervous system. Cell based therapies can restore function in spinal cord injury by replacing the lost neural tissue. These therapies also rejuvenate the existing intact neurons by facilitating remyelination and by repairing and reducing progressive tissue damage and scarring. METHODS: Autologous bone marrow stem cells were collected from the patients. 5 ml of the processed sample was injected back into the patients via lumbar puncture at L1/L2 level. The bone marrow harvesting and administration was repeated every 4 weeks 3 times (12 weeks). RESULTS: Significant improvements were noticed following the injections into the patients with the duration of injury less than 6 months. ASIA grade improvements were observed in 6 out of 10 patients. VTC and walking, at least with the support, was restored in eight patients. Bladder control and sexual functions improved in three and five patients respectively. Eight patients exhibited decreased spasticity. DISCUSSION: We believe that autologous bone marrow stem cells contributed towards the neuroplaticity and/or paracrine effect due to which we observed the considerable improvements in the conditions of the patients. CONCLUSION: This preliminary proof of patient improvement reinforces the potential of autologous bone marrow stem cell treatment in the patients suffering from Spinal Cord Injury. Although the results are encouraging further studies are needed to substantiate the claims.

Efficacy of Activated 3X Platelet-Rich Plasma in the Treatment of Androgenic Alopecia.

BACKGROUND: Platelet-rich plasma (PRP) has shown remarkable beneficial effects without any major adverse reactions in the treatment of androgenic alopecia .The growth factors in activated autologous PRP induces the proliferation of dermal papilla cells. OBJECTIVES: To investigate the clinical efficacy of Platelet Rich Plasma prepared using Merisis One Step Gel Separation Technology in treatment of androgenic alopecia. METHODS: Five patients were given autologous PRP injections on the affected area of alopecia over a period of three months at interval of two - three weeks and results were assessed. RESULTS: Three months after the treatment, the patients presented clinical improvement in the hair counts, hair thickness, hair root strength and overall alopecia. CONCLUSION: PRP appears to be a cheap, effective and promising therapy for androgenic alopecia with no major adverse effects.

Treatment of AVN Using Autologous BM Stem Cells and Activated Platelet-Derived Growth Factor Concentrates.

BACKGROUND: Avascular Necrosis (AVN) of hip is a devastating condition seen in younger individuals. It is the ischemic death of the constituents of the bone cartilage of the hip. The femoral head (FH) is the most common site for AVN. It results from interruption of the normal blood flow to the FH that fits into the hip socket. Earlier studies using autologous bone marrow stem cell concentrate injections have shown encouraging results with average success rates. The current study was designed to improve significantly the cartilage regeneration and clinical outcome. METHODS: Total of 48 patients underwent autologous bone marrow stem cell and activated platelet-rich plasma derived growth factor concentrate (PRP-GFC) therapy for early and advanced stages AVN of femoral head in a single multi-specialty center. The total treatment was divided into three phases. In the phase I, all the clinical diagnostic measurements such as magnetic resonance imaging (MRI), computed tomography (CT) etc. with respect to the AVN patients and bone marrow aspiration from posterior iliac spine from the patients were carried out. In the phase II, isolation of stem cells and preparation from the patients were performed. Subsequently, in phase III, the stem cells and PRP- GFCs were transplanted in the enrolled patients. RESULTS: Ninety three percent of the enrolled AVN patients showed marked enhancement in the hip bone joint space (more than 3mm) after combined stem cells and PRP-GFC treatment as evidenced by comparison of the pre- and post-treatment MRI data thus indicative of regeneration of cartilage. The treated patients showed significant improvement in their motor function, cartilage regrowth (3 to 10mm), and high satisfaction in the two-year follow-up. CONCLUSION: Combination of stem cell and PRP-GFC therapy has shown promising cartilage regeneration in 45 out of 48 patients of AVN. This study clearly demonstrates the safety and efficacy of this treatment. Larger numbers of patients need to be evaluated to better understand the efficacy of the combined stem cell and PRP-GFC therapy on AVN patients.

Need for In Vivo Triggering of Homeostasis to Repair Irreversible Tissue Degeneration by Stem Cells through Innate and Invasive Regeneration Processes.

The regeneration patterns are innate, inherited and evolutionarily conserved mechanisms. In every individual there are certain cells and signaling networks which work together to proliferate the desired tissue lineages to replace the dead, lost and injured counter parts. This homeostasis mechanism keeps functioning of the organ system intact. There are some tissues such as skin, gut, blood, respiratory tract, uterine endometrium, testis must perpetually renew the majority of cells. As the aging advances the turnover potentials decreases under normal circumstances, some of these respond inefficiently to regenerative pressures (eg: brain and heart) while other respond quite well. Therefore creating an optimized micro environment using external means through non-invasive or invasive procedures preferably minimally by utilizing appropriately subjected stem cells/stem cell secretome to induce regeneration at the target sites where it does not take place spontaneously.Stem cell biology is one of the most attractive areas of biomedical research, as emerges for the execution of biotechnology towards the regenerative medicine continues to expand. The presumed potential of the stem cell populations with clonogenic capabilities are harnessed for the therapeutic applications. Advancements in the research technology and the idea of inducing innate regeneration by stem cell based approaches can generate the potential cure for many degenerative disorders, age related disabilities and accidental tissue damages.

Therapeutic Application of Bone Marrow-Derived Stem Cells in a Patient with Methanol-Induced Blindness.

OBJECTIVE: To report bone marrow derived stem cell application in a patient with bilateral visual loss due to accidental ingestion of methanol. METHOD: A 22-year-old male patient with bilateral visual loss at discharge after treatment for methanol ingestion was treated with retrobulbar and intrathecal autologus bone marrow derived stem cells in an outpatient setting. The procedure was followed by Vit B12 Injection 500 mcg alternate days and oral vitamins B1, B2, B6, in therapeutic dose for 3 months. RESULT: The patient demonstrated favorable outcome three days after the treatment. Visual acuity improved to 20/20 in both eyes by third week. CONCLUSION: Treatment with retrobulbar and intrathecal autologous bone marrow derived stem cell in an adult male patient with bilateral visual loss due to accidental ingestion of methanol was effective. Larger studies are warranted to explore the real impact of this procedure.

Antiaging Effects of an Intensive Mind and Body Therapeutic Program through Enhancement of Telomerase Activity and Adult Stem Cell Counts.

OBJECTIVE: Key modalities of integrative medicine known to rejuvenate the mind and body are meditation, yoga, and controlled diet. It has been shown previously that intensive or prolonged mind and body therapies (MBT) may have beneficial effects on the well-being of healthy people and in patients. Telomerase activity and levels of peripheral blood adult pluripotent stem cells (PB-APSC) are reliable markers of long-term well-being that are known to decrease with age. The objective of this study is to understand the effect of our MBT program on telomerase activity and stem cells in blood collected from the participants. DESIGN: Here, we have investigated the effects of an intensive three weeks MBT retreat on telomerase activity and the peripheral blood stem cells in participants before and after the MBT. A total of 108 people were enrolled in the study; 38 men and 70 women (aged 18-90) randomly assigned for the study. RESULTS: Telomerase activity was greater in retreat participants at the end of the MBT retreat. About 45% of people showed more than one-fold increase of telomerase activity after our MBT program. Furthermore, about 27% of people showed more pronounced fold increase (2-fold) in telomerase activity after the MBT. In addition, a substantial percentage of people (about 90%) exhibited increased stem cell counts after the MBT. CONCLUSIONS: The data suggest increased telomerase activity and stem cells count in peripheral blood from MBT retreat participants that may lead to increased longevity and better quality of life at latter age.

Reversal of Methanol-Induced Blindness in Adults by Autologous Bone Marrow-Derived Stem Cells: A Case Series.

INTRODUCTION: Methanol ingestion leads to severe damage to visual pathways and permanent loss of vision. Current treatment is aimed at removal of methanol from system and prevention of generation of toxic metabolites along with symptomatic management of patient. Autologous bone marrow mononuclear stem cells (MNC) can be used to rejuvenate the damaged retinal cells and restoration of vision. METHODS: Five patients suffering from methanol induced complete blindness within three months of insult and no known comorbidities during the past 6 months were enrolled to receive autologous bone marrow derived mononuclear cell fraction on compassionate grounds. The visual acuity and visual evoked responses (VER) were done at the time of enrollment and during follow-up visits. OBSERVATIONS AND RESULTS: Visual acuity of these patients at the time of enrollment: no perception of light. Improvement in visual acuity was recorded by 7 days which reached maximum at 3 weeks after treatment in three patients and three months in two patients. The patients had acuity of 6/9, finger counting and reading with magnifying glasses with no subsequent improvement till 2 years of follow-up. Visual Evoked Responses demonstrated improvements following treatment. No adverse reactions were noticed during follow-up. CONCLUSION: Treatment with Autologous Bone marrow derived MNC offers a new line of management in patients with loss of vision following methanol ingestion. The efficacy and safety of this line of management needs to be evaluated in controlled clinical trials.

Ligament and tendon repair through regeneration using mesenchymal stem cells.

One of Nature's gifts to mankind is mesenchymal stem cells (MSC's). They are multipotent in nature and are present literally in every tissue. Since, they possess certain characteristics of stem cells such as self-renewal and differentiation they are known to be one of the key players in normal tissue homeostasis. This novel function of mesenchymal stem cells has been explored by scientists in the field of regenerative medicine. This review gives an insight of the various sources of mesenchymal stem cells available for tissue engineering with regard to tendon and ligament and the mechanism involved during regeneration.

Biomarker discovery and biotherapeutics applications of photosynthetic light-harvesting and bioluminescence light-emitting chromophore-protein complexes in stem cell biology and regenerative medicine.

We have since the 1970's embarked on the development of biologically derived fluorophore-protein complexes that will find applications in the communicable and non-communicable disease etiology processes and their cures. We have since then become largely successful in these endeavors along with interspersed contributions also from investigators who have generally restricted to working in confined disciplines. Their encompassment with our works as this investigator has traversed his definitely chosen and not merely a circumstantial, coincidental, or accidental step-wise multi-disciplinary scientific path from biophysics to regenerative medicine spanning these lines of investigations for last four decades have finally yielded the much necessitated disease related applied biological interventions for human benefits. Taking a cue from our early investigations and findings on which we call attention to the identification and characterization of the use of the primary light-emitting lumazine precursor of riboflavin which is 6,7-dimethyl-8-ribityl lumazine-protein complex which we had derived from the bioluminescence bacterium (Photobacterium phosphoreum) wherein it functions as a naturally occurring fluorescence light emitter (LumP). These in vivo phenomena have been a precursor to the subsequent developments in vitro. This in vivo to in vitro investigation path of ours has been also comprised among others of binding of photosynthetic light-harvesting marine dinoflagellate algae (Glenodinium sp.) derived peridinin-chlorophyll a-protein (PerCP) complex-labeled monoclonal antibodies useful in the development of flow cytometry. These fluorescence labeled antibodies bound antigens which include those of communicable infectious diseases (HIV/AIDS - env-gp160, gag-p24), non-communicable but also potential hereditary and malignant disorders (Cancer/Tumor Markers - Melan-A/Mart-1 of melanoma), normal immune response cells (Human/Mouse/species cellular MHC/TCR/CD45/CD33/CD56/CD19/CD41), and of types of stem cells (CD34/CD38/c-Mpl/Oct4/Neuropilin-1/SOX17). Such antigens have been analyzed by us and other investigators by fluorescence-activated cell sorting (FACS - cell surface and intracellular binding), confocal fluorescence microscopy, or/and immunohistochemistry, to determine qualitative and quantitative antigen expression levels and their mechanistic implications. We have followed stem cell differentiation patterns and signaling mechanisms through marker antigen-antibody binding wherein the antibodies are labeled with covalently linked fluorophore-protein complexes or fluorescence emitting chromophores. These complexes among others also have included PerCP. We are also now in the process of developing flow cytometry applications of additional visible light emitting chromophore-protein complexes through industrial collaborations. The United States Navy has long been known for interest in the estimation of changes in illumination intensity in and under the oceans to track movements of enemy ships and other naval vessels.