Histopathological analysis of the synovium in trapeziometacarpal osteoarthritis.

Dorsoradial and anterior oblique ligaments were harvested during surgery in 13 patients with symptomatic trapeziometacarpal osteoarthritis, which had been graded preoperatively by a modified Eaton-Littler radiographic grading. Ligaments, including the periligamentous synovium, were stained with S100 protein, neurotrophic receptor p75, protein gene product 9.5, calcitonin gene related peptide, acetylcholine, substance P, neuropeptide Y, noradrenaline, N-methyl-D-aspartate-receptor and Met/Leu-enkephalin. The synovium was classified as showing no, low-grade or high-grade synovitis. Free nerve endings had higher immunoreactivity for substance P than for N-methyl-D-aspartate-receptor, enkephalin and noradrenaline. The synovial stroma had less immunoreactivity for N-methyl-D-aspartate-receptor than for noradrenaline, substance P and calcitonin gene related peptide. There was no relation between the grade of osteoarthritis and the visual pain analogue scale, synovitis score, immunoreactivity of all antibodies and quantity of free nerve endings or blood vessels. Synovium in trapeziometacarpal joint osteoarthritis produces several neuromediators causing a polymodal neurogenic inflammation and which may serve as biomarkers for osteoarthritis or therapeutic targets.

Altered Innervation Pattern in Ligaments of Patients with Basal Thumb Arthritis.

Purpose The population of mechanoreceptors in patients with osteoarthritis (OA) lacks detailed characterization. In this study, we examined the distribution and type of mechanoreceptors of two principal ligaments in surgical subjects with OA of the first carpometacarpal joint (CMC1). Methods We harvested two ligaments from the CMC1 of eleven subjects undergoing complete trapeziectomy and suspension arthroplasty: the anterior oblique (AOL) and dorsal radial ligament (DRL). Ligaments were divided into proximal and distal portions, paraffin-sectioned, and analyzed using immunoflourescent triple staining microscopy. We performed statistical analyses using the Wilcoxon Rank Sum test and ANOVA with post-hoc Bonferroni and Tamhane adjustments. Results The most prevalent nerve endings in the AOL and DRL of subjects with OA were unclassifiable mechanoreceptors, which do not currently fit into a defined morphological scheme. These were found in 11/11 (100%) DRLs and 7/11 (63.6%) AOLs. No significant difference existed with respect to location within the ligament (proximal versus distal) of mechanoreceptors in OA subjects. Conclusion The distribution and type of mechanoreceptors in cadavers with no to mild OA differ from those in surgical patients with OA. Where Ruffini endings predominate in cadavers with no to mild OA, unclassifiable corpuscles predominate in surgical patients with OA. These findings suggest an alteration of the mechanoreceptor population and distribution that accompanies the development of OA. Clinical Relevance Identification of a unique type and distribution of mechanoreceptors in the CMC1 of symptomatic subjects provides preliminary evidence of altered proprioception in OA.

Chemotaxis of human induced pluripotent stem cell-derived endothelial cells.

This study examined the homing capacity of human induced pluripotent stem cell-derived endothelial cells (iPSC-ECs) and their response to chemotactic gradients of stromal derived factor-1α (SDF). We have previously shown that EC derived from murine pluripotent stem cells can home to the ischemic hindlimb of the mouse. In the current study, we were interested to understand if ECs derived from human induced pluripotent stem cells are capable of homing. The homing capacity of iPSC-ECs was assessed after systemic delivery into immunodeficient mice with unilateral hindlimb ischemia. Furthermore, the iPSC-ECs were evaluated for their expression of CXCR4 and their ability to respond to SDF chemotactic gradients in vitro. Upon systemic delivery, the iPSC-ECs transiently localized to the lungs but did not home to the ischemic limb over the course of 14 days. To understand the mechanism of the lack of homing, the expression levels of the homing receptor, CXCR4, was examined at the transcriptional and protein levels. Furthermore, their ability to migrate in response to chemokines was assessed using microfluidic and scratch assays. Unlike ECs derived from syngeneic mouse pluripotent stem cells, human iPSC-ECs do not home to the ischemic mouse hindlimb. This lack of functional homing may represent an impairment of interspecies cellular communication or a difference in the differentiation state of the human iPSC-ECs. These results may have important implications in therapeutic delivery of iPSC-ECs.

The modulation of endothelial cell morphology, function, and survival using anisotropic nanofibrillar collagen scaffolds.

Endothelial cells (ECs) are aligned longitudinally under laminar flow, whereas they are polygonal and poorly aligned in regions of disturbed flow. The unaligned ECs in disturbed flow fields manifest altered function and reduced survival that promote lesion formation. We demonstrate that the alignment of the ECs may directly influence their biology, independent of fluid flow. We developed aligned nanofibrillar collagen scaffolds that mimic the structure of collagen bundles in blood vessels, and examined the effects of these materials on EC alignment, function, and in vivo survival. ECs cultured on 30-nm diameter aligned fibrils re-organized their F-actin along the nanofibril direction, and were 50% less adhesive for monocytes than the ECs grown on randomly oriented fibrils. After EC transplantation into both subcutaneous tissue and the ischemic hindlimb, EC viability was enhanced when ECs were cultured and implanted on aligned nanofibrillar scaffolds, in contrast to non-patterned scaffolds. ECs derived from human induced pluripotent stem cells and cultured on aligned scaffolds also persisted for over 28 days, as assessed by bioluminescence imaging, when implanted in ischemic tissue. By contrast, ECs implanted on scaffolds without nanopatterning generated no detectable bioluminescent signal by day 4 in either normal or ischemic tissues. We demonstrate that 30-nm aligned nanofibrillar collagen scaffolds guide cellular organization, modulate endothelial inflammatory response, and enhance cell survival after implantation in normal and ischemic tissues.

Conversion of human fibroblasts to functional endothelial cells by defined factors.

OBJECTIVE: Transdifferentiation of fibroblasts to endothelial cells (ECs) may provide a novel therapeutic avenue for diseases, including ischemia and fibrosis. Here, we demonstrate that human fibroblasts can be transdifferentiated into functional ECs by using only 2 factors, Oct4 and Klf4, under inductive signaling conditions. APPROACH AND RESULTS: To determine whether human fibroblasts could be converted into ECs by transient expression of pluripotency factors, human neonatal fibroblasts were transduced with lentiviruses encoding Oct4 and Klf4 in the presence of soluble factors that promote the induction of an endothelial program. After 28 days, clusters of induced endothelial (iEnd) cells seemed and were isolated for further propagation and subsequent characterization. The iEnd cells resembled primary human ECs in their transcriptional signature by expressing endothelial phenotypic markers, such as CD31, vascular endothelial-cadherin, and von Willebrand Factor. Furthermore, the iEnd cells could incorporate acetylated low-density lipoprotein and form vascular structures in vitro and in vivo. When injected into the ischemic limb of mice, the iEnd cells engrafted, increased capillary density, and enhanced tissue perfusion. During the transdifferentiation process, the endogenous pluripotency network was not activated, suggesting that this process bypassed a pluripotent intermediate step. CONCLUSIONS: Pluripotent factor-induced transdifferentiation can be successfully applied for generating functional autologous ECs for therapeutic applications.

Bioluminescence imaging of stem cell-based therapeutics for vascular regeneration.

Stem cell-based therapeutics show promise for treatment of vascular diseases. However, the survival of the cells after in vivo injection into diseased tissues remains a concern. In the advent of non-invasive optical imaging techniques such as bioluminescence imaging (BLI), cell localization and survival can be easily monitored over time. This approach has recently been applied towards monitoring stem cell treatments for vascular regeneration of the coronary or peripheral arteries. In this review, we will describe the application of BLI for tracking transplanted stem cells and associating their viability with therapeutic efficacy, in preclinical disease models of vascular disease.

Endothelial cells derived from human iPSCS increase capillary density and improve perfusion in a mouse model of peripheral arterial disease.

OBJECTIVE: Stem cell therapy for angiogenesis and vascular regeneration has been investigated using adult or embryonic stem cells. In the present study, we investigated the potential of endothelial cells (ECs) derived from human induced pluripotent stem cells (hiPSCs) to promote the perfusion of ischemic tissue in a murine model of peripheral arterial disease. METHODS AND RESULTS: Endothelial differentiation was initiated by culturing hiPSCs for 14 days in differentiation media supplemented with BMP-4 and vascular endothelial growth factor. The hiPSC-ECs exhibited endothelial characteristics by forming capillary-like structures in matrigel and incorporating acetylated-LDL. They stained positively for EC markers such as KDR, CD31, CD144, and eNOS. In vitro exposure of hiPSC-ECs to hypoxia resulted in increased expression of various angiogenic related cytokines and growth factors. hiPSC-ECs were stably transduced with a double fusion construct encoded by the ubiquitin promoter, firefly luciferase for bioluminescence imaging and green fluorescence protein for fluorescent detection. The hiPSC-ECs (5×10(5)) were delivered by intramuscular injection into the ischemic hindlimb of SCID mice at day 0 and again on day 7 after femoral artery ligation (n=8). Bioluminescence imaging showed that hiPSC-ECs survived in the ischemic limb for at least 2 weeks. In addition, laser Doppler imaging showed that the ratio of blood perfusion was increased by hiPSC-EC treatment by comparison to the saline-treated group (0.58±0.12 versus 0.44±0.04; P=0.005). The total number of capillaries in the ischemic limb of mice receiving hiPSC-EC injections was greater than those in the saline-treated group (1284±155 versus 797±206 capillaries/mm(2)) (P<0.002). CONCLUSION: This study is a first step toward development of a regenerative strategy for peripheral arterial disease based on the use of ECs derived from hiPSCs.