Ether lipids influence cancer cell fate by modulating iron uptake.

Cancer cell fate has been widely ascribed to mutational changes within protein-coding genes associated with tumor suppressors and oncogenes. In contrast, the mechanisms through which the biophysical properties of membrane lipids influence cancer cell survival, dedifferentiation and metastasis have received little scrutiny. Here, we report that cancer cells endowed with a high metastatic ability and cancer stem cell-like traits employ ether lipids to maintain low membrane tension and high membrane fluidity. Using genetic approaches and lipid reconstitution assays, we show that these ether lipid-regulated biophysical properties permit non-clathrin-mediated iron endocytosis via CD44, leading directly to significant increases in intracellular redox-active iron and enhanced ferroptosis susceptibility. Using a combination of in vitro three-dimensional microvascular network systems and in vivo animal models, we show that loss of ether lipids also strongly attenuates extravasation, metastatic burden and cancer stemness. These findings illuminate a mechanism whereby ether lipids in carcinoma cells serve as key regulators of malignant progression while conferring a unique vulnerability that can be exploited for therapeutic intervention.

The effect of physical therapy and mechanical stimulation on dysfunction of lower extremities after total pelvic exenteration in cervical carcinoma patient with rectovesicovaginal fistula induced by radiotherapy: a case report.

BACKGROUND: Total pelvic exenteration is the ultimate solution for rectovesicovaginal fistula caused by radiation therapy, yet total pelvic exenteration frequently causes intraoperative complications and postoperative complications. These complications are responsible for the dysfunction of lower extremities, impaired quality of life, and even the high long-term morbidity rate, thus multidisciplinary cooperation and early intervention for prevention of complications are necessary. Physical therapy was found to reduce the postoperative complications and promote rehabilitation, yet the effect on how physiotherapy prevents and treats complications after total pelvic exenteration and pelvic lymphadenectomy remains unclear. CASE PRESENTATION: A 50-year-old Chinese woman gradually developed perianal and pelvic floor pain and discomfort, right lower limb numbness, and involuntary vaginal discharge owing to recurrence and metastasis of cervical cancer more than half a year ago. Diagnosed as rectovesicovaginal fistula caused by radiation, she received total pelvic exenteration and subsequently developed severe lower limb edema, swelling pain, obturator nerve injury, and motor dysfunction. The patient was referred to a physiotherapist who performed rehabilitation evaluation and found edema in both lower extremities, right inguinal region pain (numeric pain rate scale 5/10), decreased temperature sensation and light touch in the medial thigh of the right lower limb, decreased right hip adductor muscle strength (manual muscle test 1/5) and right hip flexor muscle strength (manual muscle test 1/5), inability actively to adduct and flex the right hip with knee extension, low de Morton mobility Index score (0/100), and low Modified Barthel Index score (35/100). Routine physiotherapy was performed in 2 weeks, including therapeutic exercises, mechanical stimulation and electrical stimulation as well as manual therapy. The outcomes showed that physiotherapy significantly reduced lower limb pain and swelling, and improved hip range of motion, motor function, and activities of daily living, but still did not prevent thrombosis. CONCLUSION: Standardized physical therapy demonstrates the effect on postoperative complications after total pelvic exenteration and pelvic lymphadenectomy. This supports the necessity of multidisciplinary cooperation and early physiotherapy intervention. Further research is needed to determine the causes of thrombosis after standardized intervention, and more randomized controlled trials are needed to investigate the efficacy of physical therapy after total pelvic exenteration.

Phosphoglycerate kinase 1 acts as a cargo adaptor to promote EGFR transport to the lysosome.

The epidermal growth factor receptor (EGFR) plays important roles in multiple cellular events, including growth, differentiation, and motility. A major mechanism of downregulating EGFR function involves its endocytic transport to the lysosome. Sorting of proteins into intracellular pathways involves cargo adaptors recognizing sorting signals on cargo proteins. A dileucine-based sorting signal has been identified previously for the sorting of endosomal EGFR to the lysosome, but a cargo adaptor that recognizes this signal remains unknown. Here, we find that phosphoglycerate kinase 1 (PGK1) is recruited to endosomal membrane upon its phosphorylation, where it binds to the dileucine sorting signal in EGFR to promote the lysosomal transport of this receptor. We also elucidate two mechanisms that act in concert to promote PGK1 recruitment to endosomal membrane, a lipid-based mechanism that involves phosphatidylinositol 4,5-bisphosphate [PI(4,5)P2] and a protein-based mechanism that involves hepatocyte growth factor receptor substrate (Hrs). These findings reveal an unexpected function for a metabolic enzyme and advance the mechanistic understanding of how EGFR is transported to the lysosome.

MiR-224-5p inhibits osteoblast differentiation and impairs bone formation by targeting Runx2 and Sp7.

Osteoporosis is a complicated multifactorial disorder characterized by low bone mass and deteriorated bone microarchitecture with an elevated fracture risk. MicroRNAs play important roles in osteoblastic differentiation. In the present study, we found that miR-224-5p was markedly downregulated during the osteogenic differentiation of C2C12 cells. Overexpression of miR-224-5p in C2C12 cells inhibited osteoblast activity, as indicated by reduced ALP activity, matrix mineralization and the expression of osteogenic marker genes. Moreover, we demonstrated that Runx2 and Sp7 were direct targets of miR-224-5p. Furthermore, the specific inhibition of miR-224-5p by femoral bone marrow cavity injection with miR-224-5p antagomir prevented ovariectomy-induced bone loss. Finally, we found that the levels of miR-224-5p were markedly elevated in the sera of patients with osteoporosis. Collectively, this study revealed that miR-224-5p negatively regulates osteogenic differentiation by targeting Runx2 and Sp7. It also highlights the potential use of miR-224-5p as a therapeutic target and diagnostic biomarker for osteoporosis. Supplementary information: The online version contains supplementary material available at 10.1007/s10616-023-00593-z.

ALKBH5-mediated m6A demethylation of HS3ST3B1-IT1 prevents osteoarthritis progression.

HS3ST3B1-IT1 was identified as a downregulated long noncoding RNA in osteoarthritic cartilage. However, its roles and mechanisms in the pathogenesis of osteoarthritis (OA) are unclear. In this study, we demonstrated that the expressions of HS3ST3B1-IT1 and its maternal gene HS3ST3B1 were downregulated and positively correlated in osteoarthritic cartilage. Overexpression of HS3ST3B1-IT1 significantly increased chondrocyte viability, inhibited chondrocyte apoptosis, and upregulated extracellular matrix (ECM) proteins, whereas HS3ST3B1-IT1 knockdown had the opposite effects. In addition, HS3ST3B1-IT1 significantly ameliorated monosodium-iodoacetate-induced OA in vivo. Mechanistically, HS3ST3B1-IT1 upregulated HS3ST3B1 expression by blocking its ubiquitination-mediated degradation. Knockdown of HS3ST3B1 reversed the effects of HS3ST3B1-IT1 on chondrocyte viability, apoptosis, and ECM metabolism. AlkB homolog 5 (ALKBH5)-mediated N6-methyladenosine (m6A) demethylation stabilized HS3ST3B1-IT1 RNA. Together, our data revealed that ALKBH5-mediated upregulation of HS3ST3B1-IT1 suppressed OA progression by elevating HS3ST3B1 expression, suggesting that HS3ST3B1-IT1/HS3ST3B1 may serve as potential therapeutic targets for OA treatment.

Targeting 14-3-3ζ by a small-molecule compound AI-34 maintains epithelial barrier integrity and alleviates colitis in mice via stabilizing ß-catenin.

Aberrant intestinal epithelial barrier function is the primary pathology of Ulcerative colitis (UC), making it a desirable drug target. In this study, our small-molecule compound AI-34 exerted a significant protective effect in an LPS-induced epithelial barrier injury model. In vitro, AI-34 treatment significantly decreased cell permeability, increased transmembrane resistance, and maintained the junctional protein (ZO-1 and E-cadherin) levels in monolayer cells. Using the LiP-small molecule mapping approach (LiP-SMap), we demonstrated that AI-34 binds to 14-3-3ζ. AI-34 promoted the interaction between 14-3-3ζ and ß-catenin, decreasing the ubiquitination of ß-catenin and thus maintaining intestinal epithelial barrier function. Finally, AI-34 triggered the stabilization of ß-catenin mediated by 14-3-3ζ, provoking a significant improvement in the DSS-induced colitis model. Our findings suggest that AI-34 may be a promising candidate for UC treatment.

METTL3-mediated m6A modification of IGFBP7-OT promotes osteoarthritis progression by regulating the DNMT1/DNMT3a-IGFBP7 axis.

Osteoarthritis (OA) is the most common degenerative disorder, affecting approximately half of the elderly population. In this study, we find that the expressions of long noncoding RNA (lncRNA) IGFBP7-OT and its maternal gene, IGFBP7, are upregulated and positively correlated in osteoarthritic cartilage. Overexpression of IGFBP7-OT significantly inhibits chondrocyte viability, promotes chondrocyte apoptosis, and reduces extracellular matrix components, whereas IGFBP7-OT knockdown has the opposite effects. IGFBP7-OT overexpression promotes cartilage degeneration and markedly aggravates the monosodium iodoacetate-induced OA phenotype in vivo. Further mechanistic research reveals that IGFBP7-OT promotes OA progression by upregulating IGFBP7 expression. Specifically, IGFBP7-OT suppresses the occupancy of DNMT1 and DNMT3a on the IGFBP7 promoter, thereby inhibiting methylation of the IGFBP7 promoter. The upregulation of IGFBP7-OT in OA is partially controlled by METTL3-mediated N6-methyladenosine (m6A) modification. Collectively, our findings reveal that m6A modification of IGFBP7-OT promotes OA progression by regulating the DNMT1/DNMT3a-IGFBP7 axis and provide a potential therapeutical target for OA treatment.

Spirodalesol analog 8A inhibits NLRP3 inflammasome activation and attenuates inflammatory disease by directly targeting adaptor protein ASC.

Pharmacological inhibition of the Nod-like receptor family protein 3 (NLRP3) inflammasome contributes to the treatment of numerous inflammation-related diseases, making it a desirable drug target. Spirodalesol, derived from the ascomycete fungus Daldinia eschscholzii, has been reported to inhibit NLRP3 inflammasome activation. Based on the structure of spirodalesol, we synthesized and screened a series of analogs to find a more potent inhibitor. Analog compound 8A was identified as the most potent selective inhibitor for NLRP3 inflammasome assembly, but 8A did not inhibit the priming phase of the inflammasome. Specifically, while 8A did not reduce NLRP3 oligomerization, we found that it inhibited the oligomerization of adaptor protein apoptosis-associated speck-like protein containing a caspase activation and recruitment domain (ASC), as ASC speck formation was significantly reduced. Also, 8A interrupted the assembly of the NLRP3 inflammasome complex and inhibited the activation of caspase-1. Subsequently, we used a cellular thermal shift assay and microscale thermophoresis assay to demonstrate that 8A interacts directly with ASC, both in vitro and ex vivo. Further, 8A alleviated lipopolysaccharide-induced endotoxemia, as well as monosodium urate-induced peritonitis and gouty arthritis in mice by suppressing NLRP3 inflammasome activation. Thus, 8A was identified as a promising ASC inhibitor to treat inflammasome-driven diseases.

m6A-mediated upregulation of AC008 promotes osteoarthritis progression through the miR-328-3p‒AQP1/ANKH axis.

Long noncoding RNAs (lncRNAs) have emerged as important regulators of osteoarthritis (OA), but the biological roles and clinical significance of most lncRNAs in OA are not fully understood. Microarray analysis was performed to identify differentially expressed lncRNAs, mRNAs, and miRNAs between normal and osteoarthritic cartilage. We found that AC008440.5 (abbreviated AC008), as well as AQP1 and ANKH, were highly expressed in osteoarthritic cartilage, whereas miR-328-3p was expressed at a low level in osteoarthritic cartilage. Functional assays showed that ectopic expression of AC008, AQP1, and ANKH significantly decreased chondrocyte viability and promoted chondrocyte apoptosis and extracellular matrix (ECM) degradation, whereas knockdown of AC008, AQP1, and ANKH resulted in the opposite effects. Moreover, miR-328-3p overexpression increased chondrocyte viability and attenuated chondrocyte apoptosis and ECM degradation, whereas inhibition of miR-328-3p resulted in the opposite effects. Bioinformatics analysis, RNA immunoprecipitation (RIP), and luciferase assays revealed that AC008 functioned as a competing endogenous RNA (ceRNA) to regulate miR-328-3p, which specifically targeted the AQP1 and ANKH genes. In addition, miR-328-3p significantly ameliorated MIA-induced OA, whereas AC008 accelerated OA progression in vivo. Furthermore, fat mass and obesity-associated (FTO)-mediated N6-methyladenosine demethylation downregulated AC008 transcription, while lower FTO expression led to upregulation of AC008 transcription in OA. In conclusion, our data reveal that AC008 plays a critical role in OA pathogenesis via the miR-328-3p‒AQP1/ANKH pathway, suggesting that AC008 may be a potential therapeutic target for OA.

GRASP55 regulates intra-Golgi localization of glycosylation enzymes to control glycosphingolipid biosynthesis.

The Golgi apparatus, the main glycosylation station of the cell, consists of a stack of discontinuous cisternae. Glycosylation enzymes are usually concentrated in one or two specific cisternae along the cis-trans axis of the organelle. How such compartmentalized localization of enzymes is achieved and how it contributes to glycosylation are not clear. Here, we show that the Golgi matrix protein GRASP55 directs the compartmentalized localization of key enzymes involved in glycosphingolipid (GSL) biosynthesis. GRASP55 binds to these enzymes and prevents their entry into COPI-based retrograde transport vesicles, thus concentrating them in the trans-Golgi. In genome-edited cells lacking GRASP55, or in cells expressing mutant enzymes without GRASP55 binding sites, these enzymes relocate to the cis-Golgi, which affects glycosphingolipid biosynthesis by changing flux across metabolic branch points. These findings reveal a mechanism by which a matrix protein regulates polarized localization of glycosylation enzymes in the Golgi and controls competition in glycan biosynthesis.

Inhibition of NLRP3 inflammasome activation in myeloid-derived suppressor cells by andrographolide sulfonate contributes to 5-FU sensitization in mice.

5-Fluorouracil (5-FU)-based chemotherapy is the first-line recommended regimen in colorectal cancer (CRC), but resistance limits its clinical application. Andrographolide sulfonate, a traditional Chinese medicine, is mainly used to treat infectious diseases. In the present study, we reported that andrographolide sulfonate could significantly inhibit the growth of transplanted CT26 colon cancer in mice and improve survival when combined with 5-FU. Furthermore, TUNEL assay and immunohistochemistry analysis of proliferating cell nuclear antigen, Ki-67 and p-STAT3 confirmed that co-treatment could inhibit tumor proliferation and promote apoptosis. In tumor tissues of groups that received 5-FU and andrographolide sulfonate, CD4+ and CD8+ T cell infiltration was increased, and the expression of IFN-γ and Granzyme B detected by immunohistochemistry and qPCR was upregulated, reflecting improved antitumor immunity. Finally, we verified that 5-FU significantly activated the NLR Family Pyrin Domain Containing 3 (NLRP3) inflammasome in myeloid-derived suppressor cells (MDSCs) and that andrographolide sulfonate reversed this process to sensitize cells to 5-FU. In summary, andrographolide sulfonate synergistically enhanced antitumor effects and improved antitumor immunity by inhibiting 5-FU-induced NLRP3 activation in MDSCs. These findings provide a novel strategy to address 5-FU resistance in the treatment of CRC.

Computational investigation of two-dimensional borides with planar octacoordinated main group elements.

In this communication, by means of first principles calculations, we searched stable two-dimensional borides with octacoordinated main group elements, and we found that only AlB4 monolayer is stable in the planar octacoordinate motif through our stability screenings, which can be used for electrocatalyzing the hydrogen evolution reaction.

C9N4 and C2N6S3 monolayers as promising anchoring materials for lithium-sulfur batteries: weakening the shuttle effect via optimizing lithium bonds.

The notorious polysulfide shuttle effect is a crucial factor responsible for the degradation of Li-S batteries. A good way to suppress the shuttle effect is to effectively anchor dissoluble lithium polysulfides (LPSs, Li2Sn) on appropriate substrates. Previous studies have revealed that Li of Li2Sn is prone to interact with the N of N-containing materials to form Li-N bonds. In this work, by means of density functional theory (DFT) computations, we explored the possibility to form Li bonds on ten different N-containing monolayers, including BN, C2N, C2N6S3, C9N4, a covalent triazine framework (CTF), g-C3N4, p-C3N4, C3N5, S-N2S, and T-N2S, by examining the adsorption behavior of Li2Sn (n = 1, 2, 3, 4, 6, 8) on these two-dimensional (2D) anchoring materials (AMs), and investigated the performance of the formed Li bonds (if any) in inhibiting the shuttle effect. By comparing and analyzing the nitrogen content, the N-containing pore size, charge transfer, and Li bonds, we found that the N content and N-containing pore size correlate with the number of Li bonds, and the formed Li-N bonds between LPSs and AMs correspond well with the adsorption energies of the LPSs. The C9N4 and C2N6S3 monolayers were identified as promising AMs in Li-S batteries. From the view of Li bonds, this work provides guidelines for designing 2D N-containing materials as anchoring materials to reduce the shuttle effect in Li-S batteries, and thus improving the performance of Li-S batteries.

MiR-664-3p suppresses osteoblast differentiation and impairs bone formation via targeting Smad4 and Osterix.

Osteoporosis is a metabolic disorder characterized by low bone mass and deteriorated microarchitecture, with an increased risk of fracture. Some miRNAs have been confirmed as potential modulators of osteoblast differentiation to maintain bone mass. Our miRNA sequencing results showed that miR-664-3p was significantly down-regulated during the osteogenic differentiation of the preosteoblast MC3T3-E1 cells. However, whether miR-664-3p has an impact on bone homeostasis remains unknown. In this study, we identified overexpression of miR-664-3p inhibited the osteoblast activity and matrix mineralization in vitro. Osteoblastic miR-664-3p transgenic mice exhibited reduced bone mass due to suppressed osteoblast function. Target prediction analysis and experimental validation confirmed Smad4 and Osterix (Osx) are the direct targets of miR-664-3p. Furthermore, specific inhibition of miR-664-3p by subperiosteal injection with miR-664-3p antagomir protected against ovariectomy-induced bone loss. In addition, miR-664-3p expression was markedly higher in the serum from patients with osteoporosis compared to that from normal subjects. Taken together, this study revealed that miR-664-3p suppressed osteogenesis and bone formation via targeting Smad4 and Osx. It also highlights the potential of miR-664-3p as a novel diagnostic and therapeutic target for osteoporotic patients.

Accurate electronic properties and non-linear optical response of two-dimensional MA2Z4.

Two-dimensional MA2Z4 (M = Mo, W, V, Nb, Ta, Ti, Zr, Hf, or Cr; A = Si or Ge; Z = N, P, or As) is a new lead in the 2D family, because it exhibits versatile properties by tuning the components M, A and Z. However, theoretical studies on MA2Z4 are quite limited, and electronic properties are mainly studied by standard DFT levels, which seriously underestimates the band gap. Here, we systematically investigated the electronic properties and nonlinear optical response of MA2Z4 using a hybrid HSE06 functional. It was found that replacing component Z changes the lattice constant most, while the lattice influence by component M substitution is only slight. We showed that the gap difference between PBE and HSE06 is generally about 30% but can be up to 101%. (MIV = Hf, Ti, or Zr)Si2N4 possesses multi-valley characteristics. Furthermore, the second-harmonic generation (SHG) responses of various MA2Z4 composites were also calculated. Three non-zero elements of second order non-linear susceptibilities are revealed for MA2Z4 with the relationship: d16 = d21 = d22, indicating that MA2Z4 belongs to the D3H1 space group. HfSi2N4 possesses a multi-valley characteristic, and exhibits the largest susceptibility under broad wavelengths and the value of d21 reaches 3697.04 pm V-1 at band gap resonance energy. Intriguingly, the non-linear coefficients of MoSi2P4 and MoSi2As4 in the IR region are two orders of magnitude larger than those of other well-known non-linear crystals, such as LiGaS2 and BaAl4S7. We further explored the anisotropic SHG response by the polar plot of intensity under different incident light into MA2Z4. Our work provides theoretical guidelines for further experimental explorations of MA2Z4 and paves the way for its utilization in non-linear optic devices.

Combined immunodeficiency due to a mutation in the γ1 subunit of the coat protein I complex.

The coat protein I (COPI) complex mediates retrograde trafficking from the Golgi to the endoplasmic reticulum (ER). Five siblings with persistent bacterial and viral infections and defective humoral and cellular immunity had a homozygous p.K652E mutation in the γ1 subunit of COPI (γ1-COP). The mutation disrupts COPI binding to the KDEL receptor and impairs the retrieval of KDEL-bearing chaperones from the Golgi to the ER. Homozygous Copg1K652E mice had increased ER stress in activated T and B cells, poor antibody responses, and normal numbers of T cells that proliferated normally, but underwent increased apoptosis upon activation. Exposure of the mutants to pet store mice caused weight loss, lymphopenia, and defective T cell proliferation that recapitulated the findings in the patients. The ER stress-relieving agent tauroursodeoxycholic acid corrected the immune defects of the mutants and reversed the phenotype they acquired following exposure to pet store mice. This study establishes the role of γ1-COP in the ER retrieval of KDEL-bearing chaperones and thereby the importance of ER homeostasis in adaptive immunity.

Long noncoding RNA FGF14-AS2 inhibits breast cancer metastasis by regulating the miR-370-3p/FGF14 axis.

Long noncoding RNAs (lncRNAs) have emerged as important regulators in cancers, including breast cancer. However, the overall biological roles and clinical significance of most lncRNAs are not fully understood. This study aimed to elucidate the potential role of a novel lncRNA FGF14-AS2 and the mechanisms underlying metastasis in breast cancer. The lncRNA FGF14-AS2 was significantly downregulated in breast cancer tissues; patients with lower FGF14-AS2 expression had advanced clinical stage. In vitro and in vivo assays of FGF14-AS2 alterations revealed a complex integrated phenotype affecting breast cancer cell migration, invasion, and tumor metastasis. Mechanistically, FGF14-AS2 functioned as a competing endogenous RNA of miR-370-3p, thereby leading to the activation of its coding counterpart, FGF14. Clinically, we observed increased miR-370-3p expression in breast cancer tissues, whereas FGF14 expression was decreased in breast cancer tissues compared to the adjacent normal breast tissues. FGF14-AS2 expression was significantly negatively correlated with miR-370-3p expression, and correlated positively to FGF14 expression. Collectively, our findings support a model in which the FGF14-AS2/miR-370-3p/FGF14 axis is a critical regulator in breast cancer metastasis, suggesting a new therapeutic direction in breast cancer.

Bovine CAPN3 core promoter initiates expression of foreign genes in skeletal muscle cells by MyoD transcriptional regulation.

Activating foreign genes in bovine skeletal muscle is necessary in the study of the role of related genes in skeletal muscle development and the effects on skeletal muscle formation, especially in the study of transgenic cattle. At this time, a skeletal muscle-specific promoter should be selected to initiate a functional foreign gene. Here, calpain3 (CAPN3) was found to be highly expressed in skeletal muscle and skeletal muscle cells by real-time PCR. Next, 5' deletion analysis of the bovine CAPN3 promoter was performed and showed that Q5(-495/+40) region was the core promoter of the bovine CAPN3. A key regulatory site (-465/-453) in CAPN3 core promoter was associated with the transcription factor, MyoD, which is a skeletal muscle-specific transcription factor. Furthermore, the mRNA and protein expression levels of MyoD and CAPN3 were positively correlated during skeletal muscle cell differentiation. The overexpression of MyoD enhanced the activity of the bovine CAPN3 core promoter. The core promoter Q5(-495/+40) could drive the exogenous gene EGFP and the fat-specific expression gene PPARγ in skeletal muscle cells. In summary, our study obtained a bovine skeletal muscle-specific promoter and provided a basis for studying the role of functional genes in the growth and development of skeletal muscle. It also provides a basis for studying the transcriptional regulation mechanism of CAPN3.

The protein kinase Akt acts as a coat adaptor in endocytic recycling.

Coat proteins have a central role in vesicular transport by binding to cargoes for their sorting into intracellular pathways. Cargo recognition is mediated by components of the coat complex known as adaptor proteins1-3. We previously showed that Arf-GAP with coil-coil, ANK repeat and PH domain-containing protein 1 (ACAP1) functions as an adaptor for a clathrin coat complex that has a function in endocytic recycling4-6. Here, we show that the protein kinase Akt acts as a co-adaptor in this complex, and is needed in conjunction with ACAP1 to bind to cargo proteins to promote their recycling. In addition to advancing the understanding of endocytic recycling, we uncover a fundamentally different function in which a kinase acts, as Akt in this case is an effector rather than a regulator in a cellular event.

Establishment and characterization of an immortalized human chondrocyte cell line.

OBJECTIVES: Following a specific number of mitotic divisions, primary chondrocytes undergo proliferative senescence, thwarting efforts to expand sufficient populations in vitro suitable to meet the needs of scientific research or medical therapies. Therefore, the human telomerase reverse transcriptase (TERT) was used to immortalize human chondrocyte and establish a cell line that escape from cellular senescence. RESULTS: The human chondrocytes were successfully immortalized by ectopic stable expression of TERT. The established TERT-Chondrocyte cell line showed robust proliferation capacity, even in late passages up to P20, and displayed little cellular senescence. Moreover, TERT-Chondrocyte cells at 20th passage showed similar chondrocyte properties to normal chondrocytes at early passages. CONCLUSIONS: Ectopic stable expression of TERT is an effective way to immortalized human chondrocyte. The immortalized chondrocytes displayed little cellular senescence, showed promise as an in vitro model to investigate osteoarthritis, and may be a promising resource for cell-based therapy for damaged cartilage.