Efficient Generation of Pancreatic Progenitor Cells from Induced Pluripotent Stem Cells Derived from a Non-Invasive and Accessible Tissue Source-The Plucked Hair Follicle.

The advent of induced pluripotent stem cell (iPSC) technology has brought about transformative advancements in regenerative medicine, offering novel avenues for disease modeling, drug testing, and cell-based therapies. Patient-specific iPSC-based treatments hold the promise of mitigating immune rejection risks. However, the intricacies and costs of producing autologous therapies present commercial challenges. The hair follicle is a multi-germ layered versatile cell source that can be harvested at any age. It is a rich source of keratinocytes, fibroblasts, multipotent stromal cells, and the newly defined Hair Follicle-Associated Pluripotent Stem Cells (HAP). It can also be obtained non-invasively and transported via regular mail channels, making it the ideal starting material for an autologous biobank. In this study, cryopreserved hair follicle-derived iPSC lines (HF-iPS) were established through integration-free vectors, encompassing a diverse cohort. These genetically stable lines exhibited robust expression of pluripotency markers, and showcased tri-lineage differentiation potential. The HF-iPSCs effectively differentiated into double-positive cKIT+/CXCR4+ definitive endoderm cells and NKX6.1+/PDX1+ pancreatic progenitor cells, affirming their pluripotent attributes. We anticipate that the use of plucked hair follicles as an accessible, non-invasive cell source to obtain patient cells, in conjunction with the use of episomal vectors for reprogramming, will improve the future generation of clinically applicable pancreatic progenitor cells for the treatment of Type I Diabetes.

Predictive Techniques for Neurotoxin Outcomes.

BACKGROUND: Botulinum-derived neurotoxins have become a substantial tool in dermatologists' armamentarium for facial/neck rejuvenation. Current literature discusses anatomical "danger zones" to avoid during neurotoxin injection to prevent brow ptosis, blepharoptosis, and lower facial ptosis. OBJECTIVE: The aim of this study was to determine whether lidocaine 1% local anesthetic can be used to predict botulinum toxin treatment outcomes and prevent adverse effects of unwanted paralysis. MATERIALS AND METHODS: One percent lidocaine was drawn up using BD ultra-fine 31 G (5/16″), 0.5-mL insulin syringes in the same quantity that would be drawn up for neurotoxin placement. The patient's face was cleansed and mapped; 0.1 mL of 1% lidocaine was injected × 5 sites in the glabella; and 3 sites were injected with 0.05 mL in the frontalis. The patient was assessed after 10 minutes. RESULTS: Improvement in frontalis and glabellar rhytides was appreciated, with noted "spocking" of the lateral brows. This technique allowed the authors to visualize the need for placement of toxin more laterally with eventual successful predictive placement for neurotoxin. CONCLUSION: This technique of using local 1% lidocaine allows the practitioner to devise a neurotoxin distribution map tailored for each patient to limit unwanted paralysis from improper neurotoxin placement.

Association of Central Nervous System Depression With Topical Brimonidine When Used for Hemostasis: A Serious Adverse Event.

Importance: Minor bleeding is the most common complication of dermatologic surgery. Topical brimonidine, 0.33%, gel has been reported for the use of hemostasis in dermatologic surgery. The safety profile and risk of systemic toxic effects when brimonidine is used topically for hemostasis is unknown. Objective: To determine the risk of systemic toxic effects of topical brimonidine, 0.33%, gel when used for hemostasis. Design, Setting, and Participants: In this case series from a private practice (Hollywood Dermatology), 2 patients presented for dermatologic procedures, complicated by persistent bleeding. Interventions: Patients were treated with 10 g of brimonidine, 0.33%, gel applied under occlusion for hemostasis. Main Outcomes and Measures: Mental status, cardiopulmonary function. Results: Both patients experienced deterioration of mental status, respiratory depression, and somnolence. Results from cardiac testing, laboratory workup, and imaging were negative for cardiac or neurologic etiology. Both patients improved in less than 24 hours. Conclusions and Relevance: Topical brimonidine, 0.33%, gel can result in systemic central nervous system toxic effects when used as a hemostatic agent. At present, it is not possible to define a quantity with which brimonidine can be used safely, nor can a safe wound size be defined. We, therefore, urge against the use of topical brimonidine as a hemostatic agent until its safety is further investigated.

Polymorphic eruption of pregnancy.

Polymorphic eruption of pregnancy (PEP), a specific dermatosis of pregnancy also known as pruritic urticarial papules and plaques of pregnancy (PUPPP), is a benign, self-limited skin disorder. Key features include an increased prevalence in primigravidas, onset in the third trimester, remission near the time of delivery, and association with multiple gestation pregnancy. The clinical features are crucial to diagnosis. Histopathology is nonspecific, and immunofluorescence studies help differentiate PEP from pemphigoid gestationis. The pathogenesis of PEP remains elusive, and relevant theories are reviewed. There are no associated maternal or fetal risks, and treatment is largely symptomatic.

Collagen type XII and versican are present in the early stages of cartilage tissue formation by both redifferentating passaged and primary chondrocytes.

Current approaches to cartilage tissue engineering require a large number of chondrocytes. Although chondrocyte numbers can be expanded in monolayer culture, the cells dedifferentiate and unless they can be redifferentiated are not optimal to use for cartilage repair. We took advantage of the differential effect of culture conditions on the ability of passaged and primary chondrocytes to form cartilage tissue to dissect out the extracellular matrix (ECM) molecules produced and accumulated in the early stages of passaged cell cartilage tissue formation as we hypothesized that passaged bovine cells that form cartilage accumulate a pericellular matrix that differs from cells that do not form cartilage. Twice passaged bovine chondrocytes (P2) (cartilage forming), or as a control primary chondrocytes (P0) (which do not generate cartilage), were cultured on three-dimensional membrane inserts in serum-free media. P2 redifferentiation was occurring during the first 8 days as indicated by increased expression of the chondrogenic genes Sox9, collagen type II, aggrecan, and COMP, suggesting that this is an appropriate time period to examine the ECM. Mass spectrometry showed that the P2 secretome (molecules released into the media) at 1 week had higher levels of collagen types I, III, and XII, and versican while type II collagen and COMP were found at higher levels in the P0 secretome. There was increased collagen synthesis and retention by P2 cells compared to P0 cells as early as 3 days of culture. Confocal microscopy showed that types XII, III, and II collagen, aggrecan, versican, and decorin were present in the ECM of P2 cells. In contrast, collagen types I, II, and III, aggrecan, and decorin were present in the ECM of P0 cells. As primary chondrocytes grown in serum-containing media, a condition that allows for the generation of cartilage tissue in vitro, also accumulate versican and collagen XII, this study suggests that these molecules may be necessary to provide a microenvironment that supports hyaline cartilage formation. Further study is required to determine if these molecules are also accumulated by passaged human chondrocytes and their role in promoting hyaline cartilage formation.

Serum- and growth-factor-free three-dimensional culture system supports cartilage tissue formation by promoting collagen synthesis via Sox9-Col2a1 interaction.

OBJECTIVE: One of the factors preventing clinical application of regenerative medicine to degenerative cartilage diseases is a suitable source of cells. Chondrocytes, the only cell type of cartilage, grown in vitro under culture conditions to expand cell numbers lose their phenotype along with the ability to generate hyaline cartilaginous tissue. In this study we determine that a serum- and growth-factor-free three-dimensional (3D) culture system restores the ability of the passaged chondrocytes to form cartilage tissue in vitro, a process that involves sox9. METHODS: Bovine articular chondrocytes were passaged twice to allow for cell number expansion (P2) and cultured at high density on 3D collagen-type-II-coated membranes in high glucose content media supplemented with insulin and dexamethasone (SF3D). The cells were characterized after monolayer expansion and following 3D culture by flow cytometry, gene expression, and histology. The early changes in signaling transduction pathways during redifferentiation were characterized. RESULTS: The P2 cells showed a progenitor-like antigen profile of 99% CD44(+) and 40% CD105(+) and a gene expression profile suggestive of interzone cells. P2 in SF3D expressed chondrogenic genes and accumulated extracellular matrix. Downregulating insulin receptor (IR) with HNMPA-(AM3) or the PI-3/AKT kinase pathway (activated by insulin treatment) with Wortmannin inhibited collagen synthesis. HNMPA-(AM3) reduced expression of Col2, Col11, and IR genes as well as Sox6 and -9. Co-immunoprecipitation and chromatin immunoprecipitation analyses of HNMPA-(AM3)-treated cells showed binding of the coactivators Sox6 and Med12 with Sox9 but reduced Sox9-Col2a1 binding. CONCLUSIONS: We describe a novel culture method that allows for increase in the number of chondrocytes and promotes hyaline-like cartilage tissue formation in part by insulin-mediated Sox9-Col2a1 binding. The suitability of the tissue generated via this approach for use in joint repair needs to be examined in vivo.

Platelet-rich plasma in the management of articular cartilage pathology: a systematic review.

OBJECTIVE: Using systematic review methodology, we endeavored to answer the following questions concerning the treatment of osteochondral pathology: (1) what pathologies have been treated in vivo with the use of platelet-rich plasma (PRP); (2) what methods of PRP preparation and delivery have been reported; (3) what assessment tools and comparison group have been used to assess its effectiveness; and (4) what are the clinical outcomes of its use. DATA SOURCES: A systematic literature search was performed of the OVID, EMBASE, and Evidence Based Medicine Reviews databases to identify all studies published up to October 2012 that assessed clinical outcomes of the use of PRP for the treatment of chondral and osteochondral pathology, excluding those including concomitant management of acute fractures or ligament reconstruction. DATA EXTRACTION: The included studies were reviewed and the following data were extracted and tabulated: study authors' year and journal, study design and level of evidence, pathology treated, methods of PRP preparation and delivery, and clinical outcome scores. DATA SYNTHESIS: Ten studies were included in the final analysis. The majority of studies assessed the use of PRP in the treatment of degenerative osteoarthritis of the knee or hip (representing 570 of a total of 662 joints). The majority of patients were treated with intra-articular injections, whereas 2 studies used PRP as an adjunct to surgical treatment. Significant improvements in joint-specific clinical scores (7 of 8 studies), general health scores (4 of 4 studies), and pain scores (4 of 6 studies) compared with baseline were reported up to 6-month follow-up, but few studies provided longer-term data. No studies reported worse scores compared with baseline at final follow-up. Three of 4 comparative studies reported significantly better clinical and/or pain scores when compared with hyaluronic acid injections at similar follow-up times. CONCLUSIONS: Currently, there is a paucity of data supporting the use of PRP for the management of focal traumatic osteochondral defects. There is limited evidence suggesting short-term clinical benefits with the use of PRP for symptomatic osteoarthritis of the knee, but the studies published to date are of poor quality and at high risk for bias. Further high-quality comparative studies with longer follow-up are needed to ascertain whether PRP is beneficial, either alone or as an adjunct to surgical procedures, in the management of articular cartilage pathology.

Use of fresh osteochondral glenoid allograft to treat posteroinferior bone loss in chronic posterior shoulder instability.

We report our experience with the use of fresh glenoid osteochondral allograft in the treatment of a chronic posttraumatic posterior subluxation of the shoulder associated with glenoid bone loss in a 54-year-old recreational football player. Based on the pathoanatomy of the lesion and availability of a bone bank providing fresh allograft, we opted for an open anatomic reconstruction using a fresh glenoid allograft. A posterior approach was used; the prepared allograft was placed in the appropriate anatomic position and fixed with 2 small fragment screws with washers. At 2-year follow-up, the clinical outcome is excellent. This procedure may represent an effective option for the treatment of chronic posterior shoulder instability due to glenoid bone loss. However, the long-term efficacy and the progression of glenohumeral osteoarthritis need to be evaluated.

Hyaline cartilage tissue is formed through the co-culture of passaged human chondrocytes and primary bovine chondrocytes.

To circumvent the problem of a sufficient number of cells for cartilage engineering, the authors previously developed a two-stage culture system to redifferentiate monolayer culture-expanded dedifferentiated human articular chondrocytes by co-culture with primary bovine chondrocytes (bP0). The aim of this study was to analyze the composition of the cartilage tissue formed in stage 1 and compare it with bP0 grown alone to determine the optimal length of the co-culture stage of the system. Biochemical data show that extracellular matrix accumulation was evident after 2 weeks of co-culture, which was 1 week behind the bP0 control culture. By 3 to 4 weeks, the amounts of accumulated proteoglycans and collagens were comparable. Expression of chondrogenic genes, Sox 9, aggrecan, and collagen type II, was also at similar levels by week 3 of culture. Immunohistochemical staining of both co-culture and control tissues showed accumulation of type II collagen, aggrecan, biglycan, decorin, and chondroitin sulfate in appropriate zonal distributions. These data indicate that co-cultured cells form cartilaginous tissue that starts to resemble that formed by bP0 after 3 weeks, suggesting that the optimal time to terminate the co-culture stage, isolate the now redifferentiated cells, and start stage 2 is just after 3 weeks.

Total hip arthroplasty to treat congenital musculoskeletal abnormalities in the juvenile Down Syndrome hip: review of literature with case.

Down Syndrome can result in musculoskeletal abnormalities of the hip at an early age. Avascular necrosis of the femoral head can occur as a result of slipped capital femoral epiphysis causing the patient a great deal of pain, limiting the ability to ambulate. Despite the benefits that this patient group can receive from the surgery, surgeons may be apprehensive to operate. It is our experience that these patients benefit greatly from arthroplasty without complication. In this report, we present a total hip replacement to treat avascular necrosis in an adolescent and address the concerns that surgeons may have in treating this patient population.

A systematic review of the use of platelet-rich plasma in sports medicine as a new treatment for tendon and ligament injuries.

OBJECTIVE: To evaluate, through a systematic review of the current literature, the evidence-based outcomes of the use of platelet-rich plasma (PRP) for the treatment of tendon and ligament injuries. DATA SOURCES: A search of English-language articles was performed in PubMed and EMBASE using keywords "PRP," "platelet plasma," and "platelet concentrate" combined with "tendon" and then "ligament" independently. The search was conducted through September 2010. STUDY SELECTION: Search was limited to in vivo studies. Nonhuman studies were excluded. Tissue engineering strategies, which included a combination of PRP with additional cell types (bone marrow), were also excluded. Articles with all levels of evidence were included. Thirteen of 32 retrieved articles respected the inclusion criteria. DATA EXTRACTION: The authors reviewed and tabulated data according to the year of study and journal, study type and level of evidence, patient demographics, method of PRP preparation, site of application, and outcomes. DATA SYNTHESIS: The selected studies focused on the application of PRP in the treatment of patellar and elbow tendinosis, Achilles tendon injuries, rotator cuff repair, and anterior cruciate ligament (ACL) reconstruction. Seven studies demonstrated favorable outcomes in tendinopathies in terms of improved pain and functional scores. In 3 studies on the use of PRP in ACL reconstruction, no statistically significant differences were seen with regard to clinical outcomes, tunnel widening, and graft integration. One study examined the systemic effects after the local PRP application for patellar and elbow tendinosis. CONCLUSIONS: Presently, PRP use in tendon and ligament injuries has several potential advantages, including faster recovery and, possibly, a reduction in recurrence, with no adverse reactions described. However, only 3 randomized clinical trials have been conducted.