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Chondroprogenitors (CPCs) have emerged as a promising cellular therapy for cartilage-related pathologies due to their inherent primed chondrogenic potential. Studies report that the addition of growth factors such as parathyroid hormone (PTH) and Bone Morphogenic Protein (BMP) enhance the chondroinducive potential in chondrocytes and mesenchymal stem cells. This study evaluated if supplementation of the standard culture medium for cell expansion with 1-34 PTH and BMP-9 would enhance the chondrogenic potential of CPCs and reduce their hypertrophic tendency.Human chondrocytes were isolated from patients undergoing total knee replacement for osteoarthritis (n = 3). Following fibronectin adhesion assay, passage 1 CPCs were divided and further expanded under three culture conditions (a) control, i.e., cells continued under standard culture conditions, (b) 1-34 PTH group, additional intermittent 6 h exposure with 1-34 PTH and (c) BMP-9 group, additional BMP-9 during culture expansion. All the groups were evaluated for population-doubling, cell cycle analysis, surface marker and gene expression for chondrogenesis, hypertrophy, multilineage differentiation and GAG (glycosaminoglycan)/DNA following chondrogenic differentiation.Concerning growth kinetics, the BMP-9 group exhibited a significantly lower S-phase and population-doubling when compared to the other two groups. Qualitative analysis for chondrogenic potential (Alcian blue, Safranin O staining and Toluidine blue for GAG) revealed that the BMP-9 group exhibited the highest uptake. The BMP-9 group also showed significantly higher COL2A1 expression than the control group, with no change in the hypertrophy marker expression.BMP-9 can potentially be used as an additive for CPCs expansion, to enhance their chondrogenic potential without affecting their low hypertrophic tendency. The mitigating effects of 1-34PTH on hypertrophy would benefit further investigation when used in combination with BMP-9 to enhance chondrogenesis whilst reducing hypertrophy.

PURPOSE: Cartilage-derived chondroprogenitors have been reported to possess the biological potential for cartilage repair. However, its inherent chondrogenic potential in pellet culture needs evaluation. In-vitro cartilage regeneration models based on pellet cultures have been employed to evaluate the chondrogenic potential of stem cells. Evaluation of the degree of differentiation routinely involves paraffin embedding, sectioning, and immunohistochemical staining of the pellet. However, since chondrogenic differentiation is commonly non-uniform, processing random sections could lead to inaccurate conclusions. The study aimed at assessing the inherent lineage bias of chondroprogenitors with and without chondrogenic induction, using a novel whole pellet processing technique. METHODS: Human chondroprogenitors (n=3) were evaluated for MSC markers and processed in pellet cultures either with stromal medium (uninduced) or chondrogenic differentiation medium (induced) for 28 days. The whole pellets and the conventional paraffin-embedded sectioned pellets were subjected to Collagen type II immunostaining and assessed using confocal laser microscopy. The staining intensities of the whole pellet were compared to the paraffin sections and revalidated using qRT-PCR for COL2A1 expression. RESULTS: Uninduced and induced pellets displayed Collagen type II in all the layers with comparable fluorescence intensities. COL2A1 expression in both pellets was comparable to confocal results. The study demonstrated that uninduced chondroprogenitors in pellet culture possess promising inherent chondrogenic potential. Confocal imaging of whole pellets displayed different degrees of chondrogenic differentiation in the entire pellet, thus its probable in-vivo behavior. CONCLUSION: The novel approach presented in this study could serve as an efficient in-vitro alternative for understanding translational application for cartilage repair.

Purpose Chondrocytes, isolated from articular cartilage, are routinely utilized in cell-based therapeutics for the treatment of cartilage pathologies. However, restoration of the biological tissue faces hindrance due to the formation of primarily fibrocartilaginous repair tissue. Chondroprogenitors have been reported to display superiority in terms of their chondrogenic potential and lesser proclivity for hypertrophy. In line with our recent results, comparing chondroprogenitors and chondrocytes, we undertook isolation of progenitors from the general pool of chondrocytes, based on surface marker expression, namely, CD166, CD34, and CD146, to eliminate off-target differentiation and generate cells of stronger chondrogenic potential. This study aimed to compare chondrocytes, chondroprogenitors, CD34−CD166+CD146+ sorted chondrocytes, and CD34−CD166+CD146− sorted chondrocytes. Methods Chondrocytes obtained from 3 human osteoarthritic knee joints were subjected to sorting, to isolate CD166+ and CD34− subsets, and then were further sorted to obtain CD146+ and CD146− cells. Chondrocytes and fibronectin adhesion-derived chondroprogenitors served as controls. Assessment parameters included reverse transcriptase polymerase chain reaction for markers of chondrogenesis and hypertrophy, trilineage differentiation, and total GAG/DNA content. Results Based on gene expression analysis, CD34−CD166+CD146+ sorted chondrocytes and chondroprogenitors displayed comparability and significantly higher chondrogenesis with a lower tendency for hypertrophy when compared to chondrocytes and CD34−CD166+CD146− sorted chondrocytes. The findings were also reiterated in multilineage potential differentiation with the 146+ subset and chondroprogenitors displaying lower calcification and chondroprogenitors displaying higher total GAG/DNA content compared to chondrocytes and 146− cells. Conclusion This unique progenitor-like population based on CD34−CD166+CD146+ sorting from chondrocytes exhibits efficient potential for cartilage repair and merits further evaluation for its therapeutic application.

Purpose In-vitro models of cartilage regeneration based on pellet cultures have been widely used to evaluate chondrogenic potential of the cell of interest and predict probable in-vivo behavior. However, pellet processing is a major challenge during handling (due to small size and possible damage to structural contour following sectioning and staining). The present study aimed to utilize human articular cartilage derived chondroprogenitors to assess if agarose-encapsulation of pellets prior to paraffin processing enable easier handling without affecting tissue morphology, glycosaminoglycan staining and immunohistochemical analysis of Collagen type II protein. Methods Passage 3 chondroprogenitors (n = 3) were evaluated for MSC markers using flow cytometry and subjected to chondrogenic differentiation as pellets cultures. Post-differentiation, the pellets were subjected to either: a) paraffin embedding, b) agarose encapsulation followed by paraffin embedding or c) agarose encapsulation followed by cryosectioning. All sections were subjected to histological staining for glycosaminoglycan uptake: Alcian blue, Safranin O (Bern score) and Toluidine blue with immunohistochemical processing for collagen type II protein deposition. Results With respect to staining and structural integrity, comparable uptake was seen in both paraffin sections and agarose embedded sections while the latter exhibited notably uniform pellets with distinct marginal demarcation. Although plain paraffin and agarose encapsulated sections demonstrated equivalent staining as represented by comparable Bern scores, glycosaminoglycan uptake, and Collagen type II deposition, cryosections exhibited significantly poor staining properties. Conclusion Agarose encapsulation of differentiated pellets prior to routine paraffin embedding, eases handling difficulties whilst maintaining structural integrity with optimal staining outcomes.

The potential for cartilage repair using articular cartilage derived chondroprogenitors has recently gained popularity due to promising results from in-vitro and in-vivo studies. Translation of results from in-vitro to a clinical setting requires a sufficient number of animal studies displaying significant positive outcomes. Thus, this systematic review comprehensively discusses the available literature (January 2000-March 2022) on animal models employing chondroprogenitors for cartilage regeneration, highlighting the results and limitations associated with their use.As per Preferred Reporting Items for Systematic Reviews and Meta-Analyses guidelines, a web-based search of PubMed and SCOPUS databases was performed for the following terminologies: "chondroprogenitors", "cartilage-progenitors", and "chondrogenic-progenitors", which yielded 528 studies. A total of 12 studies met the standardized inclusion criteria, which included chondroprogenitors derived from hyaline cartilage isolated using fibronectin adhesion assay (FAA) or migratory assay from explant cultures, further analyzing the role of chondroprogenitors using in-vivo animal models.Analysis revealed that FAA chondroprogenitors demonstrated the ability to attenuate osteoarthritis, repair chondral defects and form stable cartilage in animal models. They displayed better outcomes than bone marrow-derived mesenchymal stem cells but were comparable to chondrocytes. Migratory chondroprogenitors also demonstrated superiority to BM-MSCs in terms of higher chondrogenesis and lower hypertrophy, although a direct comparison to FAA-CPs and other cell types is warranted.Chondroprogenitors exhibit superior properties for chondrogenic repair; however, limited data on animal studies necessitates further studies to optimize their use before clinical translation for neo-cartilage formation.

Background: The current target of migraine treatment is focused on Triptans. Lasmiditan, a non-vasoconstrictive and highly selective 5HT1F receptor agonist is a novel therapeutic discovery for migraine for patients with cardiovascular (CV) risk factors or stable cardiovascular diseases and who fail to respond to the existing treatment. Objective: To identify an optimal dosing of Lasmiditan 100 mg versus 200 mg for the treatment of acute migraine attacks in adult patients with cardiovascular risk factors. Methods: Systematic searches were run in databases such as Cochrane Central Register of Controlled Trials (CENTRAL), Scopus, Google scholar, and PUBMED. Out of 83 study records identified, two studies were included for quantitative analysis. Results: There was a significant headache pain freedom at 2 h [Odds Ratio (OR): 0.77; 95% Confidence interval (CI): 0.64–0.92] and sustained pain freedom at 24 h (OR): 0.75; 95% CI: 0.61–0.93] in patients taking Lasmiditan 200 mg compared to those taking Lasmiditan 100 mg. The results were statistically insignificant for parameters like most bothersome symptoms (MBS) free at 2 h, headache relief at 2 h, disability level at 2 h, and global impression of change at 2 h. A combined analysis of these parameters showed a remarkable difference between both the groups favoring Lasmiditan 200 mg [OR: 0.88; 95% CI: 0.81–0.95]. Conclusion: An oral dosing of Lasmiditan 200 mg is ideal for the treatment of acute migraine in adult patients with CV risk factors for attaining headache pain freedom at 2 h and sustained pain freedom at 24 compared to Lasmiditan 100 mg.

Objective Chondroprogenitors have recently gained prominence due to promising results seen in in vitro and animal studies as a potential contender in cell-based therapy for cartilage repair. Lack of consensus regarding nomenclature, isolation techniques, and expansion protocols create substantial limitations for translational research, especially given the absence of distinct markers of identification. The objective of this systematic review was to identify and collate information pertaining to hyaline cartilage–derived chondroprogenitors, with regard to their isolation, culture, and outcome measures. Design As per Preferred Reporting Items for Systematic Reviews and Meta-Analyses guidelines, a web-based search of Scopus and PubMed databases was performed from January 2000 to May 2020, which yielded 509 studies. A total of 65 studies were identified that met the standardized inclusion criteria which comprised of, but was not limited to, progenitors derived from fibronectin adhesion, migrated subpopulation from explant cultures, and single-cell sorting. Result Literature search revealed that progenitors demonstrated inherent chondrogenesis and minimal tendency for hypertrophy. Multiple sources also demonstrated significantly better outcomes that bone marrow–derived mesenchymal stem cells and comparable results to chondrocytes. With regard to progenitor subgroups, collated evidence points to better and consistent outcomes with the use of migratory progenitors when compared to fibronectin adhesion assay–derived progenitors, although a direct comparison between the two cell populations is warranted. Conclusion Since chondroprogenitors exhibit favorable properties for cartilage repair, efficient characterization of progenitors is imperative, to complete their phenotypic profile, so as to optimize their use in translational research for neocartilage formation.

Objectives Accurate diagnosis of osteoarthritis (OA) is the first important step in ensuring appropriate management of the disease. A multitude of tests involving assessment of biomarkers help in assessment of severity and grading of osteoarthritic damage. However, most tests are time consuming and are limited by the paucity in synovial fluid volume. In majority of OA effusions, calcium containing crystals are found. The aim of our study was to evaluate whether a correlation existed between the amount of calcium containing crystals present in synovial fluid and severity scoring of OA to propose a quick and inexpensive technique for disease assessment. Materials and methods Monosodium-iodoacetate was used to induce high- and low-grade knee OA in adult New Zealand white rabbits (n = 6 joint each group). At 16 weeks, synovial fluid and joints were harvested for histopathological analysis. OA grading was established based on OARSI scoring. Synovial fluid calcium crystal count was assessed by light microscopy (Alizarin red) and confirmed by Fluo-4, AM imaging and polarized microscopy. Statistical analysis was performed using unpaired Student t-test and Pearson correlation. Results and conclusion The clumps counted in low-grade OA were significantly lower than high-grade OA, in addition to showing a positive correlation (coefficient: 0.65; P=0.021) between calcium crystal count and the grade of OA created. Fluo-4, AM staining, and polarized microscopy were indicative of calcium pyrophosphate dihydrate crystals. This is the first study to suggest that Alizarin red could serve as an effective and rapid, bed-side method for screening and assessing disease progression.

Depression is one of the most common turmoil encountered by pregnant women. Mothers’ suffering from antenatal depression holds a greater risk in developing postpartum depression. This increases the need for early detection of antenatal depression which could reduce further complications. Although there are several aetiologies for antenatal depression, iron deficiency anaemia (IDA) proves to be an emerging correlation. It is hypothesised that an altered myelination induced by iron deprivation dysregulates the release of neurotransmitters such as serotonin and dopamine, leading to depression. Of the four literature studies reviewed, two studies portrayed an association between IDA and antenatal depression. The remaining two literature studies showed the relation between antenatal depression and its complications in the infants. In this review article, we intend to establish an association between IDA and antenatal depression.

BACKGROUND: The ability to grow new cartilage remains the standard goal of any treatment strategy directed at cartilage repair. Chondroprogenitors have garnered interest due to their applicability in cell therapy. Pulsed electromagnetic field (PEMF) favors chondrogenesis by possible upregulation of genes belonging to TGFβ superfamily. Since TGFβ is implicated in chondrogenic signalling, the aim of the study was to evaluate the ability of PEMF to induce chondrogenesis via endogenous TGFβ production in chondroprogenitors vs differentiation using chondrogenic medium inclusive of TGFβ. METHODS: Chondroprogenitors were harvested from three non-diseased human knee joints via fibronectin assay. Passage 3 pellets were subjected to four different culture conditions: a) negative control contained chondrogenic medium without TGFβ2, b) positive control contained medium with TGFβ2, c) PEMF 1 contained medium of negative control plus single exposure to PEMF and d) PEMF 2 contained medium of negative control plus multiple exposures to PEMF. Following differentiation (day 21), pellets were assessed for gene expression of ACAN, SOX9, COL2A1, TGFβ1, TGFβ2, and TGFβ3. Alcian blue staining to detect glycosaminoglycan deposition was also performed. Medium supernatant was used to detect endogenous latent TGF-β1 levels using ELISA. RESULTS: All study arms exhibited comparable gene expression without any significant difference. Although positive control and PEMF study arms demonstrated notably better staining than negative control, the level of latent TGF-β1 was seen to be significantly high in supernatant from positive control (P

Purpose Articular chondroprogenitors, a suitable contender for cell-based therapy in cartilage repair, routinely employ fetal bovine serum (FBS) for expansion and differentiation. The possibility of transplant rejections or zoonoses transmissions raise a need for xeno-free alternatives. Use of human platelet lysate (hPL), a nutrient supplement abundant in growth factors, has not been reported for human chondroprogenitor expansion thus far. Our aim was to compare the biological profile of chondroprogenitors grown in hPL versus FBS. Methods Chondroprogenitors were isolated from 3 osteoarthritic knee joints. Following differential fibronectin adhesion assay, passage 0 cells grown in (a) 10% FBS and (b) 10% hPL were considered for assessment of growth kinetics, surface marker expression, gene expression, and trilineage differentiation. Latent transforming growth factor–β1 (TGFβ1) levels were also measured for each culture medium used. Results Cellular proliferation was significantly higher in cells grown with hPL ( P < 0.01). Surface marker expression was comparable except in CD-146 where hPL group had significantly higher values ( P = 0.03). Comparison of mRNA expression revealed notably low values of collagen I, collagen X, aggrecan, and collagen II ( P < 0.05). Trilineage differentiation was seen in both groups with higher alizarin red uptake noted in hPL. There were also significantly higher levels of latent TGFβ1 in the medium containing hPL as compared to FBS. Conclusions This is the first in vitro xeno-free study to affirm that hPL can serve as an optimal growth supplement for expansion of articular chondroprogenitors, although an in-depth assessment of resident growth factors and evaluation of different dilutions of hPL is required to assess suitability for use in translational research.

A good understanding of red cell indexes can aid medical students in a considerable manner, serving as a basis to unravel both concepts in red cell physiology and abnormalities associated with the same. In this study, we tried to assess whether an interactive animation was helpful in improving student comprehension and understanding of red cell indexes compared with conventional classroom teaching. Eighty-eight first-year MBBS students participated, of which 44 were assigned to group A and 44 were assigned to group B after randomization. After further creation of smaller groups, students were provided with 45 min to revise red cell indexes, after which they were required to complete a multimodal questionnaire. Group A subgroups used written material for revision, whereas group B subgroups had access to an interactive animation. After completion of the questionnaire, group A students also used the animation after which feedback was collected from all students. Efficacy of the animation to improve learning and retention was demonstrated, as group B students scored significantly higher than group A students on the questionnaire ( P = 0.0003). A clear majority of the students agreed/strongly agreed that the animation was easy to operate, conveyed important concepts efficiently, and improved their knowledge of related clinical aspects as well. From the results and feedback, we found that the animation was a simple, well-received model, which, by significantly improving student performance, corroborated our hypothesis that inclusion of interactive animation into student curriculum can advance their academic attainment, compared with didactic teaching alone.

IntroductionArticular cartilage is made up of hyaline tissue embodying chondrocytes, which arise from mesenchymal stromal cells (MSCs) and specialized extracellular matrix. Despite possessing resident progenitors in and around the joint primed for chondrogenesis, cartilage has limited intrinsic capacity of repair and cell turnover. Advances in isolation, culture, and characterization of these progenitors have raised the possibility for their use in cell-based cartilage repair. Chondroprogenitors (CPCs) have been classified as MSCs and have been postulated to play a vital role in injury response and are identified by their colony forming ability, proliferative potential, telomere dynamics, multipotency, and expression of stem cell markers. The combined presence of CPCs and chondrocytes within the same tissue compartments and the ability of chondrocytes to dedifferentiate and acquire stemness during culture expansion has obscured our ability to define and provide clear-cut differences between these 2 cell populations.ObjectiveThis review aims to evaluate and summarize the available literature on CPCs in terms of their origin, growth kinetics, molecular characteristics, and differential and therapeutic potential with emphasis on their difference from daughter chondrocytes.DesignFor this systematic review, a comprehensive electronic search was performed on PubMed and Google Scholar using relevant terms such as chondrocytes, chondroprogenitors, and surface marker expression.Results and ConclusionOur comparative analysis shows that there is an ill-defined distinction between CPCs and chondrocytes with respect to their cell surface expression (MSC markers and CPC-specific markers) and differentiation potential. Accumulating evidence indicates that the 2 subpopulations may be distinguished based on their growth kinetics and chondrogenic marker.

ObjectiveCell based therapy optimization is constantly underway since regeneration of genuine hyaline cartilage is under par. Although single source derivation of chondrocytes and chondroprogenitors is advantageous, lack of a characteristic differentiating marker obscures clear identification of either cell type which is essential to create a biological profile and is also required to assess cell type superiority for cartilage repair. This study was the first attempt where characterization was performed on the two cell populations derived from the same human articular cartilage samples.DesignCells obtained from normal/osteoarthritic knee joints were expanded in culture (up to passage 10). Characterization studies was performed using flow cytometry, gene expression was studied using RT-PCR, growth kinetics and tri-lineage differentiation was also studied to construct a better biological profile of chondroprogenitors as well as chondrocytes.Results and conclusionsOur results suggest that sorting based on CD34(-), CD166(+) and CD146(+), instead of isolation using fibronectin adhesion assay (based on CD49e+/CD29+), would yield a population of cells primarily composed of chondroprogenitors which when derived from normal as opposed to osteoarthritic cartilage, could provide translatable results in terms of enhanced chondrogenesis and reduced hypertrophy; both indispensable for the field of cartilage regeneration.

ObjectivesBone-marrow mesenchymal stem cells (MSCs) and chondrocytes are currently used for cell-based therapy in cartilage repair. Chondroprogenitors (CPs), resident cells of articular cartilage, demonstrate likeness to stem cells. Reports suggest that chondrocytes phenotype is altered in culture, thus making differentiation between the two cell populations difficult. Our objectives were to electrophysiologically assess chondrocytes and CPs, compare their mRNA expression with that of ionic channels already reported in MSCs, and to observe the effect of time in culture and osteoarthritic damage on cells.Design and ResultsChondrocytes and CPs at passages 0 (p0) and 5 (p5) derived from normal and osteoarthritic (OA) knee joints were used. Ionic currents were recorded by subjecting cells to depolarizing voltage pulses, and reverse transcriptase-polymerase chain reaction (RT-PCR) was used for studying ion channel expression. Our results demonstrated that both chondrocytes and CPs showed the presence of similar currents belonging to voltage-gated potassium channel subfamily, with RT-PCR confirming high mRNA expression of Maxi K, HKv1.1, HKv1.4, HKv4.2, and hEAG1 channels. Our finding also suggested that CPs were comparatively more sensitive to increased time in culture and inflammatory processes as observed in OA, as was evidenced by the significant decrease in mean current density (p0 normal CP: 183.171 ± 50.80 pA/pF; p5 normal CP: 50.225 ± 17.63 pA/pF; P = 0.0280) and significant increase in cellular size (p0 normal CP: 21.564 ± 2.98 pF; p0 OA CP: 37.939 ± 3.55 pF; P = 0.0057).ConclusionBoth cell types appear to be optimal candidates for cell-based therapy although initial seeding density, cell source (normal vs. OA), and time in culture are matters of concern, prior to cell-type selection.