Perichondrial progenitor cells promote proliferation and chondrogenesis of mature chondrocytes

Autologous chondrocytes are effective sources of cell therapy for engineering cartilage tissue to repair chondral defects, such as degenerative arthritis. The expansion of cells with chondrocyte characteristics has become a major challenge due to inadequate donor sites and poor proliferation of mature chondrocytes. The perichondrial progenitor cells (P cells) from the cambium layer of the perichondrium possessed significantly higher mesenchymal stem cell markers than chondrocytes (C cells). In the transwell co-culture system, P cells increased the passaging capacity of C cells from P6 to P9, and the cell number increased 128 times. This system increased the percentage of Alcian blue-positive chondrocytes from 40% in P6 to 62% in P9, contributing about 198 times more Alcian blue-positive chondrocytes than the control group. C cells co-cultured with P cells also exhibited higher proliferation than C cells cultured with P cell-conditioned medium. Similar results were obtained in nude mice that were subcutaneously implanted with C cells, P cells, or a mixture of the two cell types, in which the presence of both cells enhanced neocartilage formation in vivo. In aggregate, P cells enhanced the proliferation of C cells in a dose-dependent manner and prolonged the longevity of mature chondrocytes for clinical applications.

Evolution of the Cell

Cells are life’s basic building blocks, and there is no more profound question than how they came to be. What made this murky subject accessible is the invention of methods to sequence nucleic acids and proteins, and to infer evolutionary relationships from those sequences. It seems that all living things share a common ancestry in LUCA (the Last Universal Common Ancestor), a shadowy entity thought to have lived nearly 4 billion years ago. LUCA’s nature has been much debated, but she appears to have been a cell of sorts endowed with membranes, metabolic networks, a usable energy source and the machinery to express and reproduce genetic information. The earliest known event in cell history was the divergence of Archaea from Bacteria, about 3.5 billion years ago. Eukaryotic cells, more closely allied with Archaea than with Bacteria, appear much later, some 2 billion years ago. Their origin remains one of life’s mysteries, but the evidence currently favors a fusion or merger of an early archaeon with a bacterium; the latter became the ancestor of mitochondria, and played a major role in cell evolution. Eukaryotic cells of the contemporary kind emerged over hundreds of million years. Prominent events included a second instance of intracellular symbiosis, this time with a cyanobacterium, that introduced photosynthesis into the eukaryotic universe and initiated the plant lineage. Eukaryotic cells are the building blocks of all higher organisms. Just what has given the eukaryotic order an edge is yet another of life’s stubborn mysteries.

Living Cells, Lifeless Molecules

Cells are composed of molecules that are lifeless but special, because most of them occur in nature only in the context of life. They are essential to all the workings of life, and no one single class holds life’s secret: life is an emergent property of the collective of molecules, assembled into the elaborate structures called cells. Cells come in great profusion, but all are variations on just two patterns of organization: prokaryotes, small and relatively simple microbes, both Bacteria and Archaea; and eukaryotes (Eukarya), the larger and more complex cells that make up all animals, plants, and fungi. The molecules of life, for all their diversity, again fall mainly into just a handful of categories. The bulk of living matter consists of proteins, nucleic acids, carbohydrates and lipids. Biomolecules belong to chemistry, but their functions in the process of living place them in the realm of biology.

Synchronous spiking of cerebellar Purkinje cells during control of movements

The information that the brain transmits from one region to another is often viewed through the lens of firing rates. However, if the output neurons could vary the timing of their spikes with respect to each other, then through synchronization they could highlight information that may be critical for control of behavior. In the cerebellum, the computations that are performed by the cerebellar cortex are conveyed to the nuclei via inhibition. Yet, synchronous activity entrains nucleus neurons, making them fire. Does the cerebellar cortex rely on spike synchrony within populations of Purkinje cells (P-cells) to convey information to the nucleus? We recorded from multiple P-cells while marmosets performed saccadic eye movements and organized them into populations that shared a complex spike response to error. Before movement onset, P-cells transmitted information via a rate code: the simple spike firing rates predicted the direction and velocity of the impending saccade. However, during the saccade, the spikes became temporally aligned within the population, signaling when to stop the movement. Thus, the cerebellar cortex relies on spike synchronization within a population of P-cells, not individual firing rates, to convey to the nucleus when to stop a movement.

Delivery of miR-224-5p by Exosomes from Cancer-Associated Fibroblasts Potentiates Progression of Clear Cell Renal Cell Carcinoma

Objectives. Clear cell renal cell carcinoma (ccRCC) is the most common subtype of renal cell carcinoma. Cancer-associated fibroblasts (CAFs) as the primary components of cancer stroma can affect tumor progression by secreting exosomes, while exosomes are carriers for proteins, nucleic acids, and other agents that responsible for delivery of biological information. Given this, exosomes derived from CAFs are emerging as promising biomarkers in clinical cancer diagnosis. Nevertheless, their role in clear cell renal cell carcinoma (ccRCC) remains poorly understood. Methods. Here, we separated fibroblasts from ccRCC tissue, extracted exosomes, observed their morphology, and detected the expression of exosome marker proteins including Hsp70, CD9, and CD63. In the meantime, we labeled exosomes and performed coculture experiment to verify the delivery of miR-224-5p from CAFs to 769-P cells with exosomes as a carrier, so as to clarify the effect of CAF-derived exosomes on ccRCC cell malignant behaviors, as well as to discuss how miR-224-5p involves in above regulation. Results. Transmission electron microscopy was firstly applied, and it was noted that the exosomes we isolated were in normal range. Besides, Western blot also confirmed the presence of exosome marker proteins Hsp70, CD9, and CD63. Furthermore, coculture experiments were performed and the CAF-derived exosomes were observed to be able to facilitate the malignant behaviors of ccRCC cells, and the exosomal miR-224-5p could be internalized by ccRCC cells to participate in regulation of cell proliferation, migration, invasion, and apoptosis. Conclusion. To sum up, miR-224-5p can enter ccRCC cells via CAF-derived exosomes, in turn, promoting the malignant behaviors of ccRCC cells, which indicates that miR-224-5p has the potential severing as a therapeutic target for ccRCC.

Trichostatin A downregulates bromodomain and extra-terminal proteins to suppress osimertinib resistant non-small cell lung carcinoma

Background The third-generation epithelial growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs) have shown significant therapeutic effects on patients with non-small cell lung carcinoma (NSCLC) who carry active EGFR mutations, as well as those who have developed acquired resistance to the first-generation of EGFR-TKIs due to the T790M mutation. However, most patients develop drug resistance after 8–10 months of treatment. Currently, the mechanism has not been well clarified, and new therapeutic strategies are urgently needed. Methods Osimertinib resistant cell lines were established by culturing sensitive cells in chronically increasing doses of osimertinib. The anticancer effect of reagents was examined both in vitro and in vivo using the sulforhodamine B assay and a xenograft mouse model. The molecular signals were detected by western blotting. The combination effect was analyzed using CompuSyn software. Results We found that bromodomain and extra-terminal proteins (BETs) were upregulated in osimertinib resistant (H1975-OR) cells compared with those in the paired parental cells (H1975-P), and that knockdown of BETs significantly inhibited the growth of H1975-OR cells. The BET inhibitor JQ1 also exhibited stronger growth-inhibitory effects on H1975-OR cells and a greater expression of BETs and the downstream effector c-Myc than were observed in H1975-P cells. The histone deacetylase (HDAC) inhibitor trichostatin A (TSA) showed stronger growth suppression in H1975-OR cells than in H1975-P cells, but vorinostat, another HDAC inhibitor, showed equal inhibitory efficacy in both cell types. Consistently, downregulation of BET and c-Myc expression was greater with TSA than with vorinostat. TSA restrained the growth of H1975-OR and H1975-P xenograft tumors. The combination of TSA and JQ1 showed synergistic growth-inhibitory effects in parallel with decreased BET and c-Myc expression in both H1975-OR and H1975-P cells and in xenograft nude mouse models. BETs were not upregulated in osimertinib resistant HCC827 cells compared with parental cells, while TSA and vorinostat exhibited equal inhibitory effects on both cell types. Conclusion Upregulation of BETs contributed to the osimertinib resistance of H1975 cells. TSA downregulated BET expression and enhanced the growth inhibitory effect of JQ1 both in vitro and in vivo. Our findings provided new strategies for the treatment of osimertinib resistance.

AnnexinA7 and CAP1 are associated with regulating tumor cell adhesion molecule expression and biological behavior

IntroductionTo find out the correlations between the effects of down-regulating AnnexinA7 and CAP1 gene on cell adhesion factors and the biological behavior of Hca-p cells cells, and finally infer the relationship between the two genes.Material and methodsWestern blot ,qRT-PCR,immunocytochemistry, CCK8 cell proliferation, flow cytometry, lymph node adhesion and transwell chamber assay testing .ResultsAnnexinA7 and CAP1 were consistent with the regulation of the expression of the adhesion molecules such as FAK, Src, Paxillin and E-cadherin. At the mRNA and protein levels, the expression of FAK, Src and Paxillin were increased with down-regulated AnnexinA7 and CAP1 genes, while E-cadherin was down-regulated in the change of these two genes. And the low expression of AnnexinA7 could affect the expression of CAP1 in mRNA and protein levels. otherwise, the localization of AnnexinA7 and CAP1 in hepatocellular carcinoma cells was also the same. After down-regulating the expression of CAP1, the functions of proliferation, lymph node adhesion and invasion were increased and early apoptotic ability was decreased in Hca-P cells.ConclusionsAnnexinA7 and CAP1 could control the expression of these adhesion molecules in the same trend. We speculated they may be co-localization. And AnnexinA7 gene may be related to the molecular mechanism of CAP1 gene, which is likely to have a consistent effect on cell adhesion molecules and be closely related to the biological behaviors of Hca-P cells.AnnexinA7 and CAP1 may play an inhibitory role in lymph node metastasis to provide a reliable basis for the early identification of lymphatic metastasis in hepatocellular carcinoma.

Ultrastructural plasma membrane asymmetries in tension and curvature promote yeast cell fusion

Cell-cell fusion is central to the process of fertilization for sexual reproduction. This necessitates the remodeling of peri-cellular matrix or cell wall material and the merging of plasma membranes. In walled fission yeast S. pombe, the fusion of P and M cells during sexual reproduction relies on the fusion focus, an actin structure that concentrates glucanase-containing secretory vesicles for local cell wall digestion necessary for membrane fusion. Here, we present a correlative light and electron microscopy (CLEM) quantitative study of a large dataset of 3D tomograms of the fusion site, which revealed the ultrastructure of the fusion focus as an actin-containing, vesicle-dense structure excluding other organelles. Unexpectedly, the data revealed asymmetries between the two gametes: M-cells exhibit a taut and convex plasma membrane that progressively protrudes into P-cells, which exhibit a more slack, wavy plasma membrane. These asymmetries are relaxed upon plasma membrane fusion, with observations of ramified pores that may result from multiple initiations or inhomogeneous expansion. We show that P-cells have a higher exo- to endocytosis ratio than M-cells, and that local reduction in exocytosis abrogates membrane waviness and compromises cell fusion significantly more in P- than M-cells. Reciprocally, reduction of turgor pressure specifically in M-cells prevents their protrusions into P-cells and delays cell fusion. Thus, asymmetric membrane conformations, which result from differential turgor pressure and exocytosis/endocytosis ratios between mating types, favor cell-cell fusion.

P-sort: an open-source software for cerebellar neurophysiology

Analysis of electrophysiological data from Purkinje cells (P-cells) of the cerebellum presents challenges for spike detection. Complex spikes have waveforms that vary significantly from one event to the next, raising the problem of misidentification. Even when complex spikes are detected correctly, the simple spikes may belong to a different P-cell, raising the danger of misattribution. Here, we analyzed data from over 300 P-cells in marmosets, macaques, and mice, using an open-source, semi-automated software called P-sort that addresses the spike identification and attribution problems. Like other sorting software, P-sort relies on nonlinear dimensionality reduction to cluster spikes. However, it also uses the statistical relationship between simple and complex spikes to merge seemingly disparate clusters, or split a single cluster. In comparison with expert manual curation, occasionally P-sort identified significantly more complex spikes, as well as prevented misattribution of clusters. Three existing automatic sorters performed less well, particularly for identification of complex spikes. To improve development of analysis tools for the cerebellum, we provide labeled data for 313 recording sessions, as well as statistical characteristics of waveforms and firing patterns.

Simulation of a Charged Al2O3 Film As An Assisting Passivation Layer For a-Si Passivated Contact p-Type Silicon Solar Cells

In this paper, a charged Al2O3 tunneling film as an assisting for amorphous Si (a-Si) passivated contact layer is proposed and theoretically simulated for its potential application in improving a-Si passivated contact p-type (a-PC-p) solar cell. The concept is based on an Ag/n+ c-Si/p c-Si/Al2O3/p+ a-Si/Al structure. The key feature is the introduction of a charged Al2O3 layer, which facilitates the tunneling of holes through an Al2O3 insulator layer accompanied by the reduction of interface defect density (Dit). The negative charge in the Al2O3 layer makes the energy band of p-type c-Si bend upward, realizing the accumulation of holes and repelling of electrons at the c-Si/a-Si interface simultaneously. The influence of interface negative charges (Qit) between a-Si and c-Si, Al2O3 thickness, Al2O3 bandgap, interface defect density (Dit) at the a-Si/c-Si interface are systematically investigated on the output parameters of a-PC-p cells. Inserting a charged Al2O3 film between the c-Si/a-Si interface, a +4.2 % relative efficiency gain is predicted theoretically compared with the a-PC-p cells without the Al2O3 layer. Subsequently, the device performance under various temperatures is simulated, and the insertion of a charged Al2O3 layer obviously decreases the Pmax temperature coefficient from -0.336 % /℃ to -0.247 % /℃, which is analogous to that of Heterojunction with Intrinsic Thin layer (HIT) solar cell. The above results demonstrate a better temperature response for a-PC-p cells with a charged Al2O3 layer, paving a road for its potential application in high-efficiency and high thermal stability a-PC-p solar cells.