Proteomic characterization of regenerated cartilage following knee joint distraction; a human case-study.

Purpose: Knee joint distraction is a surgical procedure with cartilage-regenerating properties. The composition of joint distraction-regenerated cartilage in human patients is poorly documented. In this case-study, provided a unique opportunity to biomolecularly characterize the regenerated tissue from a patient who underwent bilateral distraction and later knee replacements.

TGF-β2 Induces Ribosome Activity, Alters Ribosome Composition and Inhibits IRES-Mediated Translation in Chondrocytes.

Alterations in cell fate are often attributed to (epigenetic) regulation of gene expression. An emerging paradigm focuses on specialized ribosomes within a cell. However, little evidence exists for the dynamic regulation of ribosome composition and function.

The BMP7-Derived Peptide p[63-82] Reduces Cartilage Degeneration in the Rat ACLT-pMMx Model for Posttraumatic Osteoarthritis.

Objective: Osteoarthritis (OA) is characterized by articular cartilage erosion, pathological subchondral bone changes, and signs of synovial inflammation and pain. We previously identified p[63-82], a bone morphogenetic protein 7 (BMP7)-derived bioactive peptide that attenuates structural cartilage degeneration in the rat medial meniscal tear-model for posttraumatic OA. This study aimed to evaluate the cartilage erosion-attenuating activity of p[63-82] in a different preclinical model for OA (anterior cruciate ligament transection-partial medial meniscectomy [anterior cruciate ligament transection (ACLT)-pMMx]).

Adaptation of the protein translational apparatus during ATDC5 chondrogenic differentiation.

Introduction: Ribosome biogenesis is integrated with many cellular processes including proliferation, differentiation and oncogenic events. Chondrogenic proliferation and differentiation require a high cellular translational capacity to facilitate cartilaginous extracellular matrix production. We here investigated the expression dynamics of factors involved in ribosome biogenesis during chondrogenic differentiation and determined whether protein translation capacity adapts to different phases of chondrogenic differentiation.

BMP7 increases protein synthesis in SW1353 cells and determines rRNA levels in a NKX3-2-dependent manner.

BMP7 is a morphogen capable of counteracting the OA chondrocyte hypertrophic phenotype via NKX3-2. NKX3-2 represses expression of RUNX2, an important transcription factor for chondrocyte hypertrophy. Since RUNX2 has previously been described as an inhibitor for 47S pre-rRNA transcription, we hypothesized that BMP7 positively influences 47S pre-rRNA transcription through NKX3-2, resulting in increased protein translational capacity.

Uncovering pathways regulating chondrogenic differentiation of CHH fibroblasts.

Mutations in the non-coding snoRNA component of mitochondrial RNA processing endoribonuclease (RMRP) are the cause of cartilage-hair hypoplasia (CHH). CHH is a rare form of metaphyseal chondrodysplasia characterized by disproportionate short stature and abnormal growth plate development. The process of chondrogenic differentiation within growth plates of long bones is vital for longitudinal bone growth.

Ribosome dysfunction in osteoarthritis.

Purpose Of Review: Translation of genetic information encoded within mRNA molecules by ribosomes into proteins is a key part of the central dogma of molecular biology. Despite the central position of the ribosome in the translation of proteins, and considering the major proteomic changes that occur in the joint during osteoarthritis development and progression, the ribosome has received very limited attention as driver of osteoarthritis pathogenesis.

Recent Findings: We provide an overview of the limited literature regarding this developing topic for the osteoarthritis field.

BMP7 reduces the fibrocartilage chondrocyte phenotype.

The fibrocartilage chondrocyte phenotype has been recognized to attribute to osteoarthritis (OA) development. These chondrocytes express genes related to unfavorable OA outcomes, emphasizing its importance in OA pathology. BMP7 is being explored as a potential disease-modifying molecule and attenuates the chondrocyte hypertrophic phenotype.

Current Practice in Bicistronic IRES Reporter Use: A Systematic Review.

Bicistronic reporter assays have been instrumental for transgene expression, understanding of internal ribosomal entry site (IRES) translation, and identification of novel cap-independent translational elements (CITE). We observed a large methodological variability in the use of bicistronic reporter assays and data presentation or normalization procedures. Therefore, we systematically searched the literature for bicistronic IRES reporter studies and analyzed methodological details, data visualization, and normalization procedures.

Prediction of the Effect of the Osteoarthritic Joint Microenvironment on Cartilage Repair.

Osteoarthritis (OA) is characterized by progressive articular cartilage loss. Human mesenchymal stromal cells (MSCs) can be used for cartilage repair therapies based on their potential to differentiate into chondrocytes. However, the joint microenvironment is a major determinant of the success of MSC-based cartilage formation.

Isolation of Nucleus Pulposus and Annulus Fibrosus Cells from the Intervertebral Disc.

Cells isolated from the intervertebral disc are often used for in vitro experimentation. Correctly separating the intervertebral disc tissue in annulus fibrosus and nucleus pulposus is particularly challenging when working with surplus material from surgery or specimens from donors with an advanced age. Moreover, lineage controls are only sparsely reported to verify tissue of origin.

The antiviral protein viperin regulates chondrogenic differentiation via CXCL10 protein secretion.

Viperin (also known as radical SAM domain-containing 2 (RSAD2)) is an interferon-inducible and evolutionary conserved protein that participates in the cell's innate immune response against a number of viruses. mRNA is a substrate for endoribonucleolytic cleavage by RNase mitochondrial RNA processing (MRP) and mutations in the RNase MRP small nucleolar RNA (snoRNA) subunit of the RNase MRP complex cause cartilage-hair hypoplasia (CHH), a human developmental condition characterized by metaphyseal chondrodysplasia and severe dwarfism. It is unknown how CHH-pathogenic mutations in RNase MRP snoRNA interfere with skeletal development, and aberrant processing of RNase MRP substrate RNAs is thought to be involved.

Identification of TGFβ-related genes regulated in murine osteoarthritis and chondrocyte hypertrophy by comparison of multiple microarray datasets.

Objective: Osteoarthritis (OA) is a joint disease characterized by progressive degeneration of articular cartilage. Some features of OA, including chondrocyte hypertrophy and focal calcification of articular cartilage, resemble the endochondral ossification processes. Alterations in transforming growth factor β (TGFβ) signaling have been associated with OA as well as with chondrocyte hypertrophy.

A Membranome-Centered Approach Defines Novel Biomarkers for Cellular Subtypes in the Intervertebral Disc.

Objective: Lack of specific marker-sets prohibits definition and functional distinction of cellular subtypes in the intervertebral disc (IVD), such as those from the annulus fibrosus (AF) and the nucleus pulposus (NP).

Design: We recently generated immortalized cell lines from human NP and AF tissues; these comprise a set of functionally distinct clonal subtypes. Whole transcriptome analyses were performed of 12 phenotypically distinct clonal cell lines (4× NP-Responder, 4× NP-nonResponder, 2× AF-Sheet forming, and 2× AF-nonSheet forming).

Transcriptional profiling distinguishes inner and outer annulus fibrosus from nucleus pulposus in the bovine intervertebral disc.

Background: Cells in the intervertebral disc have unique phenotypes and marker genes that separate the nucleus pulposus (NP), annulus fibrosus (AF) and articular cartilage (AC) have been identified. Recently, it was shown that phenotypic marker genes exhibit variable expression in humans. In this study, the bovine tail was used to determine the ability of marker genes to distinguish the outer and inner AF from NP tissue and isolated cells.

SMAD3 and SMAD4 have a more dominant role than SMAD2 in TGFβ-induced chondrogenic differentiation of bone marrow-derived mesenchymal stem cells.

To improve cartilage formation by bone marrow-derived mesenchymal stem cells (BMSCs), the signaling mechanism governing chondrogenic differentiation requires better understanding. We previously showed that the transforming growth factor-β (TGFβ) receptor ALK5 is crucial for chondrogenesis induced by TGFβ. ALK5 phosphorylates SMAD2 and SMAD3 proteins, which then form complexes with SMAD4 to regulate gene transcription.

Genetic modification of ER-Hoxb8 osteoclast precursors using CRISPR/Cas9 as a novel way to allow studies on osteoclast biology.

Osteoclasts are cells specialized in bone resorption. Currently, studies on murine osteoclasts are primarily performed on bone marrow-derived cells with the use of many animals and limited cells available. ER-Hoxb8 cells are conditionally immortalized monocyte/macrophage murine progenitor cells, recently described to be able to differentiate toward functional osteoclasts.

EGR1 controls divergent cellular responses of distinctive nucleus pulposus cell types.

Background: Immediate early genes (IEGs) encode transcription factors which serve as first line response modules to altered conditions and mediate appropriate cell responses. The immediate early response gene EGR1 is involved in physiological adaptation of numerous different cell types. We have previously shown a role for EGR1 in controlling processes supporting chondrogenic differentiation.

Novel Immortal Cell Lines Support Cellular Heterogeneity in the Human Annulus Fibrosus.

Introduction: Loss of annulus fibrosus (AF) integrity predisposes to disc herniation and is associated with IVD degeneration. Successful implementation of biomedical intervention therapy requires in-depth knowledge of IVD cell biology. We recently generated unique clonal human nucleus pulposus (NP) cell lines.

Novel immortal human cell lines reveal subpopulations in the nucleus pulposus.

Introduction: Relatively little is known about cellular subpopulations in the mature nucleus pulposus (NP). Detailed understanding of the ontogenetic, cellular and molecular characteristics of functional intervertebral disc (IVD) cell populations is pivotal to the successful development of cell replacement therapies and IVD regeneration. In this study, we aimed to investigate whether phenotypically distinct clonal cell lines representing different subpopulations in the human NP could be generated using immortalization strategies.

The immediate early gene product EGR1 and polycomb group proteins interact in epigenetic programming during chondrogenesis.

Initiation of and progression through chondrogenesis is driven by changes in the cellular microenvironment. At the onset of chondrogenesis, resting mesenchymal stem cells are mobilized in vivo and a complex, step-wise chondrogenic differentiation program is initiated. Differentiation requires coordinated transcriptomic reprogramming and increased progenitor proliferation; both processes require chromatin remodeling.