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We have used multiple sequencing approaches to sequence the genome of a volunteer from Saudi Arabia. We use the resulting data to generate ade novoassembly of the genome, and use different computational approaches to refine the assembly. As a consequence, we provide a contiguous assembly of the complete genome of an individual from Saudi Arabia for all chromosomes except chromosome Y, and label this assemblyKSA001. We transferred genome annotations from reference genomes and predicted genome features using methods from Artificial Intelligence to fully annotateKSA001, and we make all primary sequencing data, the assembly, and the genome annotations freely available in public databases using the FAIR data principles.

Cancer stem cells (CSCs) use their stemness properties to perpetuate their lineage and survive chemotherapy. Currently cell-based and cell-free therapies are under investigation to develop novel anti-cancer treatment modalities. We designed this study to investigate how cell extracts of mesenchymal stem cells affect the growth of glioma stem cells in vitro. Gliospheres were generated from the U87MG cell line and treated with conditioned media of Wharton’s jelly and bone marrow mesenchymal stem cells. The effects were investigated at the functional and molecular levels. Our results showed that conditioned media from both types of mesenchymal stem cells changed the morphology of spheres and inhibited the proliferation, invasion, and self-renewal ability of glioma stem cells. At the molecular level, metabolism interruption at oxidative phosphorylation, cell cycle arrest, cell differentiation, and upregulation of the immune response were observed. Furthermore, this effect was mediated by the upregulation of the DKK1 gene inhibiting the Wnt pathway mediated by growth factor activity and downregulation of the KITLG gene activated by growth factor and cytokine activity, inhibiting multiple pathways. We conclude that different types of mesenchymal stem cells possess antitumor properties and their paracrine factors, in combination with anti-immune modalities, can provide practical therapeutic targets for glioblastoma treatment., Graphical Abstract, Glioblastoma has limited treatment options due to the selective blood-brain barrier. Paracrine factors from mesenchymal stem cells can cross this barrier and exhibited anti-oncogenic potential for glioma stem cells in vitro. These soluble factors can be a tool as a combinatorial therapy for the treatment of glioblastomas.

We report about rationally designed ultrashort peptide bioinks, overcoming severe limitations in current bioprinting procedures. Bioprinting is increasingly relevant in tissue engineering, regenerative and personalized medicine due to its ability to fabricate complex tissue scaffolds through an automated deposition process. Printing stable large-scale constructs with high shape fidelity and enabling long-term cell survival are major challenges that most existing bioinks are unable to solve. Additionally, they require chemical or UV-cross-linking for the structure-solidifying process which compromises the encapsulated cells, resulting in restricted structure complexity and low cell viability. Using ultrashort peptide bioinks as ideal bodylike but synthetic material, we demonstrate an instant solidifying cell-embedding printing process via a sophisticated extrusion procedure under true physiological conditions and at cost-effective low bioink concentrations. Our printed large-scale cell constructs and the chondrogenic differentiation of printed mesenchymal stem cells point to the strong potential of the peptide bioinks for automated complex tissue fabrication.

The stemness in keratinocyte stem cells (KSCs) is determined by their gene expression patterns. KSCs are crucial in maintaining epidermal homeostasis and wound repair and are widely used candidates for therapeutic applications. Although several studies have reported their positive identifiers, unique biomarkers for KSCs remain elusive. Here, we aim to identify potential candidate stem cell markers. Human epidermal keratinocytes (HEKs) from neonatal foreskin tissues were isolated and cultured. Single-cell clonal analysis identified and characterized three types of cells: KSCs (holoclones), transient amplifying cells (TACs, meroclones), and differentiated cells (DSCs, paraclones). The clonogenic potential of KSCs demonstrated the highest proliferation potential of KSCs, followed by TACs and DSCs, respectively. Whole-transcriptome analysis using microarray technology unraveled the molecular signatures of these cells. These results were validated by quantitative real-time polymerase chain reaction and flow cytometry analysis. A total of 301 signature upregulated and 149 downregulated differentially expressed genes (DEGs) were identified in the KSCs, compared to TACs and DSCs. Furthermore, DEG analyses revealed new sets of genes related to cell proliferation, cell adhesion, surface makers, and regulatory factors. In conclusion, this study provides a useful source of information for the identification of potential SC-specific candidate markers.

Aim: Variations in the clinical outcomes using mesenchymal stem cells (MSCs) treatments exist, reflecting different origins and niches. To date, there is no consensus on the best source of MSCs most suitable to treat a specific disease. Methods: Total transcriptome analysis of human MSCs was performed. MSCs were isolated from two adult sources bone marrow, adipose tissue and two perinatal sources umbilical cord and placenta. Results: Each MSCs type possessed a unique expression pattern that reflects an advantage in terms of their potential therapeutic use. Advantages in immune modulation, neurogenesis and other aspects were found. Discussion: This study is a milestone for evidence-based choice of the type of MSCs used in the treatment of diseases.

The therapeutic potential of mesenchymal stem cells (MSCs) for various malignancies is currently under investigation due to their unique properties. However, many discrepancies regarding their anti-tumoral or pro-tumoral properties have raised uncertainty about their application for anti-cancer therapies. To investigate, if the anti-tumoral or pro-tumoral properties are subjective to the type of MSCs under different experimental conditions we set out these experiments. Three treatments namely cell lysates (CL), serum-free conditioned media and FBS conditioned media (FBSCM) from each of Wharton’s Jelly MSCs and Bone Marrow-MSCs were applied to evaluate the anti-tumoral or pro-tumoral effect on the glioma cells (U87MG). The functional analysis included; Morphological evaluation, proliferation and migration potential, cell cycle analysis, and apoptosis for glioma cells. The fibroblast cell line was added to investigate the stimulatory or inhibitory effect of treatments on the proliferation of the normal cell. We found that cell lysates induced a generalized inhibitory effect on the proliferation of the glioma cells and the fibroblasts from both types of MSCs. Similarly, both types of conditioned media from two types of MSCs exerted the same inhibitory effect on the proliferation of the glioma cells. However, the effect of two types of conditioned media on the proliferation of fibroblasts was stimulatory from BM-MSCs and variable from WJ-MSCs. Moreover, all three treatments exerted a likewise inhibitory effect on the migration potential of the glioma cells. Furthermore, we found that the cell cycle was arrested significantly at the G1 phase after treating cells with conditioned media which may have led to inhibit the proliferative and migratory abilities of the glioma cells (U87MG). We conclude that cell extracts of MSCs in the form of secretome can induce specific anti-tumoral properties in serum-free conditions for the glioma cells particularly the WJ-MSCs and the effect is mediated by the cell cycle arrest at the G1 phase.

Supplemental Digital Content is available in the text, Introduction: People with multiple sclerosis (PwMS) experience a wide range of disabilities which negatively impact their quality of life (QOL). Several interventions have been used in PwMS such as medication, physical therapy exercises and stem cell therapy to improve their QOL. However, there is a limited evidence on the benefits of combining interventions. The purpose of this study is to explore the effect of combining physical therapy exercises (PTE) and Wharton Jelly mesenchymal stem cell (WJ-MSCs) injections on motor and non-motor symptoms versus each intervention alone in PwMS. Methods: Sixty PwMS will be allocated to either PTE, WJ-MSCs, or a combined group, followed up for 12 months and examined using a comprehensive battery of measures. Participants in the PTE group will receive 2 sessions per week of a supervised exercise program for 6 months followed by a home exercise program for another 6 months. The WJ-MSCs group will receive 3 WJ-MSCs injections in the first 6 months then they will be encouraged to follow an active life style. The third group will receive both interventions. Discussion: This study will aid in a better understanding of the combined effect of physical therapy and mesenchymal stem cell therapy. The results from this proposed study may reduce disability, improve QOL in PwMS, and consequently, reduce the cost associated with the life-time care of these individuals worldwide. Trial registration number: NCT03326505

The therapeutic potential of mesenchymal stem cells (MSCs) for various malignancies is currently under investigation due to their unique properties. However, many discrepancies regarding their anti-tumoral or pro-tumoral properties have raised uncertainty about their application for anti-cancer therapies. To investigate, if the anti-tumoral or pro-tumoral properties are subjective to the type of MSCs under different experimental conditions we set out these experiments. Three treatments namely cell lysates (CL), serum-free conditioned media and FBS conditioned media (FBSCM) from each of Wharton’s Jelly MSCs and Bone Marrow-MSCs were applied to evaluate the anti-tumoral or pro-tumoral effect on the glioma cells (U87MG). The functional analysis included; Morphological evaluation, proliferation and migration potential, cell cycle analysis, and apoptosis for glioma cells. The fibroblast cell line was added to investigate the stimulatory or inhibitory effect of treatments on the proliferation of the normal cell. We found that cell lysates induced a generalized inhibitory effect on the proliferation of the glioma cells and the fibroblasts from both types of MSCs. Similarly, both types of conditioned media from two types of MSCs exerted the same inhibitory effect on the proliferation of the glioma cells. However, the effect of two types of conditioned media on the proliferation of fibroblasts was stimulatory from BM-MSCs and variable from WJ-MSCs. Moreover, all three treatments exerted a likewise inhibitory effect on the migration potential of the glioma cells. Furthermore, we found that the cell cycle was arrested significantly at the G1 phase after treating cells with conditioned media which may have led to inhibit the proliferative and migratory abilities of the glioma cells (U87MG). We conclude that cell extracts of MSCs in the form of secretome can induce specific anti-tumoral properties in serum-free conditions for the glioma cells particularly the WJ-MSCs and the effect is mediated by the cell cycle arrest at the G1 phase.

Long-standing foot ulcers present a great challenge in diabetes care. Platelet products have been suggested as a possible therapeutic option. However, nor the effect of an injectable form of platelet lysate on the healing of ulcers nor that on primary cells of the epidermis have been studied. In the current study, we present two cases of an ongoing clinical trial showing the positive effect of autologous platelet lysate injected perilesional. Both clinical cases treated with injections of hPL showed complete healing of previously un-healed within 8 weeks of treatment. Further, we describe the in vitro effect of human platelet lysate (hPL) on primary human epidermal keratinocytes (HEK) in terms of chemotaxis, migration and proliferation. In vitro, HEK showed enhanced chemotaxis towards the hPL compared to keratinocyte-defined media (p < 0.0001). Their migration was also stimulated especially at hPL concentration of 10%V/V (p < 0.0001). In contrast, hPL significantly inhibited HEK proliferation measured through MTT assay (p < 0.0001). In conclusion, the findings presented here provide preliminary evidence of an explanatory mechanism for the effect of hPL on primary keratinocytes and therefore of their potential use in a clinical setting. hPL promotes keratinocyte migration and therefore closure of foot ulcers., Cell biology; Proteins; Biomedical engineering; Diabetes; Regenerative medicine; Platelet lysate; Primary keratinocytes; Migration; Wound healing

Background Knee osteoarthritis (KOA) is a major health problem especially in the aging population. There is a need for safe treatment that restores the cartilage and reduces the symptoms. The use of stem cells is emerging as a possible option for the moderate and severe cases. This study aimed at testing the safety of autologous bone marrow mesenchymal stem cells (BM-MSCs) expanded in vitro when given intra-articularly to patients with stage II and III KOA. As a secondary end point, the study tested the ability of these cells to relieve symptoms and restore the knee cartilage in these patients as judged by normalized knee injury and Osteoarthritis Outcome Score (KOOS) and by magnetic resonance imaging (MRI). Methods Thirteen patients with a mean age of 50 years suffering from KOA stages II and III were given two doses of BM-MSCs 1 month apart totaling 61 × 106 ± 0.6 × 106 by intra-articular injection in a phase I prospective clinical trial. Each patient was followed for a minimum of 24 months for any adverse events and for clinical outcome using normalized KOOS. Cartilage thickness was assessed by quantitative MRI T2 at 12 months of follow-up. Results No severe adverse events were reported up to 24 months follow-up. Normalized KOOS improved significantly. Mean knee cartilage thickness measured by MRI improved significantly. Conclusion BM-MSCs given intra-articularly are safe in knee osteoarthrosis. Despite the limited number of patients in this study, the procedure described significantly improved the KOOS and knee cartilage thickness, indicating that they may enhance the functional outcome as well as the structural component. Trial registration ClinicalTrials.gov, NCT02118519

To study the use of autologous platelet lysate prepared in a standardized method for the healing of persistent corneal epithelial defects (PED). Clinical and experimental investigation. In this prospective pilot study (ClinicalTrials.gov identifier NCT02979912), ten patients with a PED duration of a minimum 14 days were included. Autologous platelet lysate was prepared in a standardized methodology. Repeated freeze–thaw cycles were used to lyse the platelets. Patients were advised to apply the eye drops four times a day and were evaluated at baseline and on days 7, 14, 21, 28. No adverse events were reported due to the use of undiluted autologous platelet lysate. A total of 70% of patients had complete re-epithelialization within 28 days. Of these, 40% healed within 14 days (effective group) and 30% within 28 days (partially effective group). Undiluted autologous platelet lysate, prepared according to a standardized methodology, is a safe and effective adjunct therapy for the treatment of PED.

Background & Aim Background The ability of mesenchymal stem cells (MSCs) to interact with immune cells and exert a modulatory effect is the core of cellular therapy approaches. Dendritic cells (DCs), the most potent of antigen presenting cells (APCs), play a key role in T-cell activation and firing immune responses. The direct interaction between MSCs and mature DCs (mDCs) results in attenuation of mDCs' activity and subsequent immune reactions. Aim Bone marrow derived MSCs (BM-MSCs) and adipose tissue derived MSCs (AT- MSCs), are more abundant and easier to harvest compared to other types of MSCs. How BM-MSCs and AT-MSCs affect mDCs can be illustrated with changes in surface marker expression and mDCs secreted cytokines, a focus is needed on changes in mDC gene expression which may help in a better understanding of the interaction process. Methods, Results & Conclusion Methods MSCs were isolated from two adult sources bone marrow and adipose tissue. Monocytes were isolated from peripheral blood to generate mDCs in recommended culture conditions. All cells were characterized by flowcytometry. Multiplex immunoassay was carried out to evaluate the inflammatory cytokines levels. PCR arrays were performed to demonstrate the gene expression profile of mDCs. All tests were performed before and after direct contact co-culture of mDCs with BM-MSCs, and with AT-MSCs. Results The immune-inhibitory effect of both types of MSCs on mDCs was manifested by a decrease in the levels of inflammatory cytokines, and the downregulation of 84 investigated genes responsible for mDC activity in both co-cultures. The fold changes, and fold change ratios from PCR array data clearly demonstrate the superiority of AT-MSCs in inhibiting mDCs immune awakening functions. This finding is supported by previous microarray analysis of the inherent immune-modulatory potential of both types of MSCs, which revealed an advantage for AT-MSCs over BM-MSCs, in terms of the expression magnitude of immune-regulatory genes. Functional annotation clustering and gene ontology demonstrated that all investigated genes participate in response to stimulus (p- value= 4.3 E-26), and that 79.8% (67 out of 84 genes) are significant players in positive regulation of response to stimulus (p-value = 4.3 E-45). Conclusion These findings highlight the outcome of MSCs’ interaction with mDCs on the molecular level. Our data confirm the immune-modulatory properties of MSCs and suggest AT-MSCs as a more effective suppressor of mDCs immune activity than BM-MSCs.

Background & Aim Background Multiple sclerosis (MS) is the most frequent chronic inflammatory, demyelinating and neurodegenerative disease of the Central Nervous System (CNS) in young adults. The use of Autologous Mesenchymal stem cells (MSCs) have been investigated to treat MS for the past decades .Outcomes of non-standardized clinical trials have been inconsistent and calls for the use of allogenic MSCs have recently increased due to suggestions that the disease is affecting the therapeutic aspects of MSCs. Aim This work aims to investigate the influence of the genetic makeup of MS patients, the environmental factors involved in the etiology of MS or the immunomodulatory drugs taken by patients on their Bone Marrow derived Mesenchymal Stem Cells (MS-MSCs) compared to healthy donors, Control MSCs (CTR-MSCs). MSCs characteristics and immune- modulatory therapeutic potential was analyzed. Methods, Results & Conclusion Methods MSCs from 8 MS patients and 3 controls were characterized using microscopy, flow cytometry of surface markers, tri-lineage inherent and induced differentiation potential. For a comprehensive analysis on the transcriptome level, Affymetrix HTA microarray chips were used. In addition, Multiplex Enzyme Linked Immunosorbent Assay (ELISA) immunoassay to assess the Cytokine and chemokine secretion by MSCs was performed using the 27-plex human inflammatory kit (Biorad). Furthermore, coculture experiments of MS-MSCs from with T cells and DCs were conducted. Results MS-MSCs passed all the MScs characterization criteria. Microarray Analysis reveals the differential expression of two disease associated markers in MS-MSCs; miRNA-599 and the signal transducing gene T-cell activation RhoGTPase activating protein TAGAP. On the other hand, MS-MSCs showed an upregulation of important immunoregulatory genes in MS-MSCs such as TGFb1, the potent Treg differentiation factor. Moreover, IL-10 and other anti-inflammatory cytokines’ levels were higher in co-cultures of T cells with MS-MSCs. While, mDCs decreased in co-cultures with MS-MSCs. Conclusion The MS disease-related or medication-related cellular and biochemical changes in the bone marrow microenvironment does not appear to have a negative impact on the immune- modulation potential of BM-MSCs. On the contrary, there seem to have a positive conditioning effect of the inflammatory milieu which enhances the modulation of both the innate and the adaptive immunity.

Summary. Haemophilia A (HA) is an X-linked recessive bleeding disorder caused by mutations in the factor VIII gene (F8), which encodes factor VIII (FVIII) protein, a plasma glycoprotein, that plays an important role in the blood coagulation cascade. In the present study, our aim was to identify F8 gene mutations in HA patients from Jordan. One hundred and seventy-five HA patients from 42 unrelated families were included in this study. Among these patients, 117 (67%) had severe HA, 13 (7%) had moderate HA and 45 (26%) had mild HA. Severe patients were first tested for intron-22 inversion using long range polymerase chain reaction (PCR), then negative patients were tested for intron-1 inversion using PCR. Sequencing for the entire F8 gene was performed for all severe HA patients who were found negative for intron-22 and -1 inversions and it was also performed for moderate and mild HA patients. HA causative mutations were identified in all patients. Intron-22 and -1 inversions were detected in 52% and 2% of families respectively. Beside these two mutations, 19 different mutations were identified, which include 15 missense and four frameshift mutations. Five novel mutations were identified including one frameshift and four missense mutations. No large deletions or nonsense mutations were detected in patients who participated in this study. Only 17 patients with severe HA were found positive for FVIII inhibitors. The data presented will play an important role for genetic counselling and health care of HA patients in Jordan.