Morphology Expression Research Articles

Multidisciplinary high resolution Geophysical Imaging of Pantano Ripa Rossa Segment of the Irpinia Fault (Southern Italy)

The Irpinia Fault, also known as the Monte Marzano Fault System, located in the Southern Apennines (Italy), is one of the most seismically active structures in the Mediterranean. It is the source of the 1980, Ms 6.9, multi-segment rupture earthquake that caused significant damage and nearly 3,000 casualties. Paleoseismological surveys indicate that this structure has generated at least four Mw ~ 7 surface-rupturing earthquakes in the past 2 ka. This paper presents a comprehensive, high-resolution geophysical investigation focused on the southernmost fault segment of the Monte Marzano Fault System, i.e., the Pantano-Ripa Rossa Fault, outcropping within the Pantano di San Gregorio Magno intramontane basin. The project, named TEst Site IRpinia fAult (TESIRA), was supported by the University of Napoli Federico II to study the near-surface structure of this intra-basin fault splay that repeatedly ruptured co-seismically in the past thousands of years. Our imaging approach included 2D and 3D electrical and seismic surveys, gravimetry, 3D FullWaver electrical tomography, drone-borne GPR and magnetic surveys, and CO2 soil flux assessment across the surface rupture. This multidisciplinary investigation improved our understanding of the basin shallow structure, providing an image of a rather complex subsurface fault and basin geometry. Seismic data suggest that fault activity at the Pantano segment of MMFS is characterized by a near-surface cumulative displacement greater than previous estimations, calling into question earlier assumptions about the timing of its activation. Despite some challenges with our drone-mounted survey equipment, the integrated dataset provides a comprehensive and reliable image of the subsurface structure. This work demonstrates the utility of developing an integrated approach at high-resolution geophysical imaging and interpretation of fault zones with weak morphological expressions.

Mechanical confinement matters: Unveiling the effect of two-photon polymerized 2.5D and 3D microarchitectures on neuronal YAP expression and neurite outgrowth

The effect of mechanical cues on cellular behaviour has been reported in multiple studies so far, and a specific aspect of interest is the role of mechanotransductive proteins in neuronal development. Among these, yes-associated protein (YAP) is responsible for multiple functions in neuronal development such as neuronal progenitor cells migration and differentiation while myocardin-related transcription factor A (MRTFA) facilitates neurite outgrowth and axonal pathfinding. Both proteins have indirectly intertwined fates via their signalling pathways. There is little literature investigating the roles of YAP and MRTFA in vitro concerning neurite outgrowth in mechanically confined microenvironments. Moreover, our understanding of their relationship in immature neurons cultured within engineered confined microenvironments is still lacking. In this study, we fabricated, via two-photon polymerization (2PP), 2.5D microgrooves and 3D polymeric microchannels, with a diameter range from 5 to 30 μm. We cultured SH-SY5Y cells and differentiated them into immature neuron-like cells on both 2.5D and 3D microstructures to investigate the effect of mechanical confinement on cell morphology and protein expression. In 2.5D microgrooves, both YAP and MRTFA nuclear/cytoplasmic (N/C) ratios exhibited maxima in the 10 μm grooves indicating a strong relation with mechanical-stress-inducing confinement. In 3D microchannels, both proteins’ N/C ratio exhibited minima in presence of 5 or 10 μm channels, a behaviour that was opposite to the ones observed in the 2.5D microgrooves and that indicates how the geometry and mechanical confinement of 3D microenvironments are unique compared to 2.5D ones due to focal adhesion, actin, and nuclear polarization. Further, especially in presence of 2.5D microgrooves, cells featured an inversely proportional relationship between YAP N/C ratio and the average neurite length. Finally, we also cultured human induced pluripotent stem cells (hiPSCs) and differentiated them into cortical neurons on the microstructures for up to 2 weeks. Interestingly, YAP and MRTFA N/C ratios also showed a maximum around the 10 μm 2.5D microgrooves, indicating the physiological relevance of our study. Our results elucidate the possible differences induced by 2.5D and 3D confining microenvironments in neuronal development and paves the way for understanding the intricate interplay between mechanotransductive proteins and their effect on neural cell fate within engineered cell microenvironments.

Evolution: Repeat adaptation in the hot spring

Live-bearing fish have repeatedly adapted to life in sulfidic hot springs. A new study finds consistent changes in morphology, physiology and gene expression but no repeated genomic adaptation. This raises further questions about genetic redundancy, polygenic adaptation and the broader significance of repeated adaptation in natural systems.

Fatty acid composition and anti-cancer activity of essential oil from Tenebrio molitor larvae in combination with zoledronic acid on prostate cancer

The essential oil extracted from Tenebrio molitor larvae (EOTM) is a natural product containing trace elements with potential therapeutic properties. This study aimed to assess the anticancer effects of EOTM and its synergistic interactions with zoledronic acid, a bisphosphonate drug, on prostate cancer cell lines. The chemical composition of EOTM was analyzed using GC-MS revealing a high concentration of fatty acids. The cytotoxicity of EOTM, both as a standalone treatment and in combination with zoledronic acid, was evaluated on prostate cancer cell lines (LNCaP, PC3) and normal hSKM using MTT assays. Results demonstrated that EOTM exhibited selective toxicity, inhibiting the growth of cancer cells in a dose-dependent manner while sparing normal cells. Morphological assessments and gene expression analyses of BCL2 and BAX were conducted through microscopy, Western blotting, and real-time RT-qPCR. These analyses indicated that EOTM induced apoptosis in cancer cells, as evidenced by cellular shrinkage, membrane blebbing, and nuclear fragmentation. Western blot results showed that EOTM downregulated the anti-apoptotic protein BCL2 and upregulated the proapoptotic protein BAX, suggesting activation of apoptosis pathways. Additionally, the combination of EOTM with zoledronic acid amplified these effects. Hoechst 33258 staining further confirmed the purity of cells following treatment. In conclusion, EOTM exhibits strong anticancer properties by inducing apoptosis in prostate cancer cells and demonstrates synergistic potential when combined with zoledronic acid. These findings warrant further investigation of EOTM as a natural and effective cancer treatment option.

Chemomechanical regulation of EZH2 localization controls epithelial-mesenchymal transition.

The methyltransferase enhancer of zeste homolog 2 (EZH2) regulates gene expression and aberrant EZH2 expression and signaling can drive fibrosis and cancer. However, it is not clear how chemical and mechanical signals are integrated to regulate EZH2 and gene expression. We show that culture of cells on stiff matrices in concert with transforming growth factor (TGF)-β1 promotes nuclear localization of EZH2 and an increase in the levels of the corresponding histone modification, H3K27me3, thereby regulating gene expression. EZH2 activity and expression are required for TGFβ1- and stiffness-induced increases in H3K27me3 levels as well as for morphological and gene expression changes associated with epithelial-mesenchymal transition (EMT). Inhibition of Rho associated kinase (ROCK) or myosin II signaling attenuates TGFβ1-induced nuclear localization of EZH2 and decreases H3K27me3 levels in cells cultured on stiff substrata, suggesting that cellular contractility, in concert with a major cancer signaling regulator TGFβ1, modulates EZH2 subcellular localization. These findings provide a contractility-dependent mechanism by which matrix stiffness and TGFβ1 together mediate EZH2 signaling to promote EMT.

Heterogeneity of Endothelial Cells Impacts the Functionality of Human Pancreatic In Vitro Models.

Endothelial cells (ECs) play a crucial role in maintaining tissue homeostasis and functionality. Depending on their tissue of origin, ECs can be highly heterogeneous regarding their morphology, gene and protein expression, functionality, and signaling pathways. Understanding the interaction between organ-specific ECs and their surrounding tissue is therefore critical when investigating tissue homeostasis, disease development, and progression. In vitro models often lack organ-specific ECs, potentially limiting the translatability and validity of the obtained results. The goal of this study was to assess the differences between commonly used EC sources in tissue engineering applications, including human umbilical vein ECs (HUVECs), human dermal microvascular ECs (hdmvECs), and human foreskin microvascular ECs (hfmvECs), and organ-specific human pancreatic microvascular ECs (hpmvECs), and test their impact on functionality within an in vitro pancreas test system used for diabetes research. Utilizing high-resolution Raman microspectroscopy and Raman imaging in combination with established protein and gene expression analyses and exposure to defined physical signals within microfluidic cultures, we identified that ECs exhibit significant differences in their biochemical composition, relevant protein expression, angiogenic potential, and response to the application of mechanical shear stress. Proof-of-concept results showed that the coculture of isolated human islets of Langerhans with hpmvECs significantly increased the functionality when compared with control islets and islets cocultured with HUVECs. Our study demonstrates that the choice of EC type significantly impacts the experimental results, which needs to be considered when implementing ECs into in vitro models.

Dynamic culture of cerebral organoids using a pillar/perfusion plate for the assessment of developmental neurotoxicity

Despite the potential toxicity of commercial chemicals to the development of the nervous system (known as developmental neurotoxicity or DNT), conventionalin vitrocell models have primarily been employed for the assessment of acute neuronal toxicity. On the other hand, animal models used for the assessment of DNT are not physiologically relevant due to the heterogenic difference between humans and animals. In addition, animal models are low-throughput, time-consuming, expensive, and ethically questionable. Recently, human brain organoids have emerged as a promising alternative to assess the detrimental effects of chemicals on the developing brain. However, conventional organoid culture systems have several technical limitations including low throughput, lack of reproducibility, insufficient maturity of organoids, and the formation of the necrotic core due to limited diffusion of nutrients and oxygen. To address these issues and establish predictive DNT models, cerebral organoids were differentiated in a dynamic condition in a unique pillar/perfusion plate, which were exposed to test compounds to evaluate DNT potential. The pillar/perfusion plate facilitated uniform, dynamic culture of cerebral organoids with improved proliferation and maturity by rapid, bidirectional flow generated on a digital rocker. Day 9 cerebral organoids in the pillar/perfusion plate were exposed to ascorbic acid (DNT negative) and methylmercury (DNT positive) in a dynamic condition for 1 and 3 weeks, and changes in organoid morphology and neural gene expression were measured to determine DNT potential. As expected, ascorbic acid did not induce any changes in organoid morphology and neural gene expression. However, exposure of day 9 cerebral organoids to methylmercury resulted in significant changes in organoid morphology and neural gene expression. Interestingly, methylmercury did not induce adverse changes in cerebral organoids in a static condition, thus highlighting the importance of dynamic organoid culture in DNT assessment.

The effect of extracellular potassium concentration on the oscillation frequency of the pacemaker nucleus in the weakly electric fish Apteronotus leptorhynchus.

The weakly electric brown ghost knifefish (Apteronotus leptorhynchus) exhibits a pronounced sexual dimorphism in its electric behavior-males discharge at higher frequencies than females, with little overlap between the sexes. The frequency of these electric organ discharges is controlled by the frequency of the synchronized oscillations of the medullary pacemaker nucleus. Previous studies have suggested that sex-specific differences in the morphology and gene expression pattern of the astrocytic syncytium that envelopes the pacemaking neural network cause differences in its capacity to buffer the extracellular concentration of K+. This change in the K+ buffering capacity affects the K+ equilibrium potential of the neurons constituting the neural network, which in turn modulates the frequency of the pacemaker nucleus. In the present study, we have tested a critical element of this hypothesis by examining whether, and how, changes in the extracellular K+ concentration influence the frequency of the pacemaker nucleus oscillations. By using an in vitro preparation of the pacemaker nucleus, the results of this investigation demonstrate that exposure of this nucleus to acutely increased/decreased concentrations of K+ in the perfusate (while maintaining osmolarity) leads to concentration-dependent increases/decreases in the frequency of the synchronized oscillations generated by the pacemaker nucleus.

Downregulation of heat shock protein 47 caused lysosomal dysfunction leading to excessive chondrocyte apoptosis

Heat shock protein 47 (HSP47) is a collagen-specific chaperone present in several regions of the endoplasmic reticulum and cytoplasm. Elevated HSP47 expression in cells causes various cancers and fibrotic disorders. However, the consequences of HSP47 downregulation leading to chondrocyte death, as well as the underlying pathways, remain largely unclear. This study presents the first experimental evidence of the localization of HSP47 on lysosomes. Additionally, it successfully designed and generated shRNA HSP47 target sequences to suppress the expression of HSP47 in ATDC5 chondrocytes using lentiviral vectors. By employing a chondrocyte model that has undergone stable downregulation of HSP47, we observed that HSP47 downregulation in chondrocytes, disturbs the acidic homeostatic environment of chondrocyte lysosomes, causes hydrolytic enzyme activity dysregulation, impairs the lysosome-mediated autophagy-lysosome pathway, and causes abnormal expression of lysosomal morphology, number, and functional effector proteins. This implies the significance of the presence of HSP47 in maintaining proper lysosomal function. Significantly, the inhibitor CA-074 Me, which can restore the dysfunction of lysosomes, successfully reversed the negative effects of HSP47 on the autophagy-lysosomal pathway and partially reduced the occurrence of excessive cell death in chondrocytes. This suggests that maintaining proper lysosomal function is crucial for preventing HSP47-induced apoptosis in chondrocytes. The existence of HSP47 is crucial for preserving optimal lysosomal function and autophagic flux, while also inhibiting excessive apoptosis in ATDC5 chondrocytes.

Oviduct epithelial spheroids during in vitro culture of bovine embryos mitigate oxidative stress, improve blastocyst quality and change the embryonic transcriptome

BackgroundIn vitro embryo production is increasingly used for genetic improvement in cattle but bypasses the oviduct environment and exposes the embryos to oxidative stress with deleterious effects on further development. Here we aimed to examine the effect of oviduct epithelial spheroids (OES) on embryo development and quality in terms of morphology and gene expression during two co-culture times (4 days: up to embryonic genome activation at 8–16 cell stage vs. 7 days: up to blastocyst stage) and under two oxygen levels (5% vs. 20%).MethodsBovine presumptive zygotes produced by in vitro fertilization (day 0) using in-vitro matured oocytes were cultured in droplets of synthetic oviductal fluid (SOF) medium with or without (controls) OES for 4 or 7 days under 5% or 20% oxygen (4 treated and 2 control groups). Cleavage rates were evaluated on day 2 and blastocyst rates on days 7–8. Expanded blastocysts on days 7–8 were evaluated for total cell numbers and gene expression analysis by RNA-sequencing.ResultsUnder 20% oxygen, blastocyst rates and total cell numbers were significantly higher in the presence of OES for 4 and 7 days compared to controls (P < 0.05), with no difference according to the co-culture time. Under 5% oxygen, the presence of OES did not affect blastocyst rates but increased the number of cells per blastocyst after 7 days of co-culture (P < 0.05). Both oxygen level and OES co-culture had a significant impact on the embryonic transcriptome. The highest number of differentially expressed genes (DEGs) was identified after 7 days of co-culture under 20% oxygen. DEGs were involved in a wide range of functions, including lipid metabolism, membrane organization, response to external signals, early embryo development, and transport of small molecules among the most significantly impacted.ConclusionOES had beneficial effects on embryo development and quality under both 5% and 20% oxygen, mitigating oxidative stress. Stronger effects on embryo quality and transcriptome were obtained after 7 than 4 days of co-culture. This study shows the impact of OES on embryo development and reveals potential molecular targets of OES-embryo dialog involved in response to stress and early embryonic development.

Impaired pericyte-Müller glia interaction via PDGFRβ suppression aggravates photoreceptor loss in a rodent model of light-induced retinal injury.

To investigate the involvement of pericyte-Müller glia interaction in retinal damage repair and assess the influence of suppressing the platelet-derived growth factor receptor β (PDGFRβ) signaling pathway in retinal pericytes on photoreceptor loss and Müller glial response. Sprague-Dawley rats were exposed to intense light to induce retinal injury. Neutralizing antibody against PDGFRβ were deployed to block the signaling pathway in retinal pericytes through intravitreal injection. Retinal histology and Müller glial reaction were assessed following light injury. In vitro, normal and PDGFRβ-blocked retinal pericytes were cocultured with Müller cell line (rMC-1) to examine morphological and protein expression changes upon supplementation with light-injured supernatants of homogenized retinas (SHRs). PDGFRβ blockage 24h prior to intense light exposure resulted in a significant exacerbation of photoreceptor loss. The upregulation of GFAP and p-STAT3, observed after intense light exposure, was significantly inhibited in the PDGFRβ blockage group. Further upregulation of cytokines monocyte chemoattractant protein 1 (MCP-1) and interleukin-1β (IL-1β) was also observed following PDGFRβ inhibition. In the in vitro coculture system, the addition of light-injured SHRs induced pericyte deformation and upregulation of proliferating cell nuclear antigen (PCNA) expression, while Müller cells exhibited neuron-like morphology and expressed Nestin. However, PDGFRβ blockage in retinal pericytes abolished these cellular responses to light-induced damage, consistent with the in vivo PDGFRβ blockage findings. Pericyte-Müller glia interaction plays a potential role in the endogenous repair process of retinal injury. Impairment of this interaction exacerbates photoreceptor degeneration in light-induced retinal injury.

Altered Reactive Oxygen Species Scavenging and Hormonal Signaling in Tetraploid Rice Are Associated with Blast Resistance.

Autotetraploid rice shows distinct morphological, physiological, hormonal, and gene expression changes that enhance its resistance to rice blast.

Primary Mast Cell Sarcoma of the Maxillary Sinus and Gingiva Mimicking Malignant Neuroendocrine Tumor: A Case Report

Mast cell sarcoma (MCS) is an extremely rare and aggressive malignancy primarily affecting bones, with limited literature associating it with neuroendocrine marker expression. This report presents a rare case of MCS arising in the maxillary sinus and gingiva. A 74-year-old man presented with a progressively enlarging ulcer on the right-sided upper gingiva. Magnetic resonance imaging revealed a 3.4 cm tumor on the floor of the right maxillary sinus. The patient underwent an inferior maxillectomy and right-sided neck dissection. Microscopically, the tumor consisted of monotonous round cells with oval nuclei, vesicular chromatin, inconspicuous nucleoli, and brisk mitoses. A panel of immunohistochemical stains was initially applied to exclude common sinonasal undifferentiated neoplasms, such as sinonasal undifferentiated carcinoma, melanoma, rhabdomyosarcoma, Ewing sarcoma, and lymphoma. The tumor cells showed patchy staining for INSM1 and synaptophysin, but were negative for AE1/AE3, CAM5.2, p40, chromogranin, S100, HMB45, NKX2.2, desmin, CD45 (LCA), CD3, and CD20, with intact INI1 and BRG1 expression. No specific diagnosis could be rendered based on the staining results, leading to consideration of other rare malignancies. Additional staining revealed positivity for CD117, mast cell tryptase, CD13, CD33, CD43, and CD68, confirming the MCS diagnosis. Molecular testing for KIT mutation was negative. Subsequent bone marrow biopsy demonstrated infiltration of atypical mast cells, which led to a diagnosis of mast cell leukemia. Despite high-dose chemotherapy, the patient died three months after the initial diagnosis. The undifferentiated epithelioid morphology and unusual aberrant neuroendocrine marker expression posed significant diagnostic challenges. The major differential diagnoses were discussed in this report.

Both the Positioned Supplemental or Night-Interruptional Blue Light and the Age of Leaves (or Tissues) Are Important for Flowering and Vegetative Growth in Chrysanthemum.

In this study, the effects of supplemental or night interruptional blue light (S-BL or NI-BL) positioning on morphological growth, photoperiodic flowering, and expression of floral genes in Chrysanthemum morifolium were investigated. Blue light-emitting diodes (LEDs) at an intensity of 30 μmol·m-2·s-1 photosynthetic photon flux density (PPFD) were used for 4 h either (1) to supplement the white LEDs at the end of the 10 h short-day (SD10 + S-BL4) and 13 h long-day conditions (LD13 + S-BL4), or (2) to provide night interruption in the SD10 (SD10 + NI-BL4) and LD13 (LD13 + NI-BL4). The S-BL4 or NI-BL4 was positioned to illuminate either the shoot tip, the youngest leaf (vigorously growing the third leaf from the shoot tip), or the old leaf (the third leaf from the stem base). In the text, they will be denoted as follows: SD10 + S-BL4-S, -Y, or -O; SD10 + NI-BL4-S, -Y, or -O; LD13 + S-BL4-S, -Y, or -O; LD13 + NI-BL4-S, -Y, or -O. Normally, the LD13 conditions enhanced more vegetative growth than the SD10 periods. The growth of leaves, stems, and branches strongly responded to the S-BL4 or NI-BL4 when it was targeted onto the shoot tip, followed by the youngest leaf. The SD10 + S-BL4 or +NI-BL4 on the old leaf obviously suppressed plant extension growth, resulting in the smallest plant height. Under LD13 conditions, the flowering-related traits were significantly affected when the S-BL4 or NI-BL4 was shed onto the youngest leaf. However, these differences do not exist in the SD10 environments. At the harvest stage, other than the non-flowered LD13 treatment, the LD13 + S-BL4 irradiating the youngest leaf induced the most flowers, followed by the shoot tip and old leaf. Moreover, LD13 + NI-BL4 resulted in the latest flowering, especially when applied to the shoot tip and old leaf. However, the SD10 + S-BL4 or + NI-BL4 irradiated the shoot tip, youngest leaf, or old leaf all significantly earlier and increased flowering compared to the SD10 treatment. Overall: (1) Generally, vegetative growth was more sensitive to photoperiod rather than lighting position, while, during the same photoperiod, the promotion of growth was stronger when the light position of S-BL4 or NI-BL4 was applied to the shoot tip or the youngest leaf. (2) The photoperiodic flowering of these short-day plants (SDPs) comprehensively responded to the photoperiod combined with blue light positioning. Peculiarly, when they were exposed to the LD13 flowering-inhibited environments, the S-BL4 or NI-BL4 shed onto the leaves, especially the youngest leaves, significantly affecting flowering.

Thyroidectomy Induced Toxicity of Testis and Possible Amelioration by Chia (Salvia hispanica) Seeds Extract

Background: Thyroidectomy or thyroid surgery induces hypothyroidism and causes thyroid and testicular hormone deficiency. The current Study Aimed to examine the therapeutic effect of chia seeds extract against thyroidectomy-induced testicular toxicity, oxidative stress, DNA damage, proliferation in rats. Methods: 40 male rats were equally divided into 4 groups [Gp1, control ; Gp2, CSE ; Gp3, thyroidectomies rats; Gp4, treated thyroidectomies rats with chia seeds extract (Thy+CSE )]. Result: Following a thyroidectomy, significant reductions (P less than 0.05) were observed in The Levels of triiodothyronine (T3), thyroxine (T4), sexual hormones (Testosterone (tt), Follicle-Stimulating Hormone and Luteinizing Hormone And prolactin), count of sperm, index of morphology, vitality, progressive motility, total motility (PR+NP), Glutathione, Superoxide Dismutase, Glutathione S-Transferase and PCNA expressions compared to the control and CSE after three weeks. Additionally, thyroidectomy leads to significant (P less than 0.05) increases in the serum of thyrotropin (TSH), alanine transaminase, alkaline phosphatase, non-progressive sperms, immotile sperms, malondialdehyde, DNA damage and injury of testis and significant (P less than 0.05) decreases in blood. Thy+CSE treatment helped to modulate most of these changes and recover testicular injury when compared To The Control and CSE. Rats treated with CSE after thyroidectomy Showed gradual improvements in testicular functions, histology and spermatogenesis due to its beneficial affection in free radicals’ scavenger.

Extracellular vesicles derived from Msh homeobox 1 (Msx1)-overexpressing mesenchymal stem cells improve digit tip regeneration in an amputee mice model

In adult mammals, limb regeneration is limited by the absence of blastemal cells (BCs) and the lack of the regenerative signaling cascade. The utilization of transgenic cells circumvents the limitations associated with the absence of BCs. In a previous investigation, we successfully regenerated mouse phalanx amputations using blastema-like cells (BlCs) generated from bone marrow-derived mesenchymal stem cells (mBMSCs) overexpressing Msx1 and Msx2 genes. Recently, extracellular vesicles (EVs) have emerged as potent biological tools, offering a promising alternative to manipulated cells for clinical applications. This research focuses on utilizing BlCs-derived extracellular vesicles (BlCs-EVs) for regenerating mouse digit tips. The BlCs were cultured and expanded, and then EVs were isolated via ultracentrifugation. The size, morphology, and CD81 marker expression of the EVs were confirmed through Dynamic Light Scattering (DLS), Scanning Electron Microscope (SEM), and Western Blot (WB) analyses. Additionally, WB analysis demonstrated the presence of MSX1, MSX2, FGF8, and BMP4 proteins. The uptake of EVs by mBMSCs was shown through immunostaining. Effects on cell proliferation, migration, and osteogenic activity post-treatment with BlCs-EVs were assessed through MTT assay, scratch assay, and Real-time PCR. The regenerative potential of BlCs-EVs was evaluated in a mouse digit tip amputation model using histological assessments. Results indicated that BlCs-EVs enhanced several abilities of mBMSCs, such as migration, proliferation, and osteogenesis in vitro. Notably, BlCs-EVs significantly improved digit tip regeneration in mice, promoting the formation of new bone and nails, which was absent in control groups. In summary, BlCs-EVs are promising tools for digit tip regeneration, avoiding the ethical concerns associated with using genetically modified cells.

A network of transient domains for breaking symmetry during anterior-posterior axis formation in the porcine embryo.

Breaking radial symmetry for anterior-posterior axis formation is one of the key developmental steps of vertebrate gastrulation and is established through a succession of transient domains defined by morphology or gene expression. Three such domains were interpreted recently in the rabbit to be part of a "three-anchor-point model" for axis formation. To answer the question as to whether the model is generally applicable to mammals, the dynamic expression patterns of four marker genes were analyzed in the pig, where gastrulating epiblast forms from half the inner cell mass: EOMES and PKDCC transcripts display decreasing expression intensities in the anterior hypoblast and-together with WNT3-increasing intensity in the anterior streak domain and the node; TBX6 expression changes from an initial central expression to exclusive expression in the posterior extremity of the primitive streak. The anterior streak domain has thus a molecular footprint similar to the one in the rabbit, the end node shares TBX6 between the species, while the anterior hypoblast-mirroring specific porcine epiblast derivation and fate-is marked by PKDCC instead of WNT3. The molecular similarities in transient domains point to conserved mechanisms for establishing the mammalian anterior-posterior axis and, possibly, breaking radial symmetry.

Maternal stress and the early embryonic microenvironment: investigating long-term cortisol effects on bovine oviductal epithelial cells using air–liquid interface culture

The oviduct epithelium is the initial maternal contact site for embryos after fertilization, offering the microenvironment before implantation. This early gestation period is particularly sensitive to stress, which can cause reduced fertility and reproductive disorders in mammals. Nevertheless, the local impact of elevated stress hormones on the oviduct epithelium has received limited attention to date, except for a few reports on polyovulatory species like mice and pigs. In this study, we focused on the effects of chronic maternal stress on cattle, given its association with infertility issues in this monoovulatory species. Bovine oviduct epithelial cells (BOEC) differentiated at the air–liquid interface (ALI) were stimulated with 250 nmol/L cortisol for 1 or 3 weeks. Subsequently, they were assessed for morphology, bioelectrical properties, and gene expression related to oviduct function, glucocorticoid pathway, cortisol metabolism, inflammation, and apoptosis. Results revealed adverse effects of cortisol on epithelium structure, featured by deciliation, vacuole formation, and multilayering. Additionally, cortisol exposure led to an increase in transepithelial potential difference, downregulated mRNA expression of the major glucocorticoid receptor (NR3C1), upregulated the expression of cortisol-responsive genes (FKBP5, TSC22D3), and significant downregulation of oviductal glycoprotein 1 (OVGP1) and steroid receptors PGR and ESR1. The systematic comparison to a similar experiment previously performed by us in porcine oviduct epithelial cells, indicated that bovine cultures were more susceptible to elevated cortisol levels than porcine. The distinct responses between both species are likely linked to their divergence in the cortisol-induced expression changes of HSD11B2, an enzyme controlling the cellular capacity to metabolise cortisol. These findings provide insights into the species-specific reactions and reproductive consequences triggered by maternal stress.

The Optejet Technology Minimizes Preservative-Mediated Cytotoxicity of Conjunctival Epithelial Cells Treated with Latanoprost In Vitro.

Purpose: Benzalkonium chloride (BAK) is a commonly used preservative to maintain sterility for multiuse eye drops such as latanoprost. One option to minimize the deleterious effects of BAK in eye drops may be to reduce the volume administered. The aim of this study was to assess the response of cells from the ocular surface to latanoprost+BAK administered by the Optejet technology, which dispenses a microdose (∼8 µL) ophthalmical spray. Methods: Cultured human conjunctival epithelial cells were exposed to the following treatments: (1) no treatment, (2) drop form of latanoprost without BAK (∼35 µL), (3) drop form of latanoprost with 0.01% BAK (∼35 µL), (4) ophthalmical spray form of latanoprost with 0.01% BAK delivered by the Optejet technology (∼8 µL). After 5 h, cells were assessed for changes in cytotoxicity, morphology, and inflammatory marker expression. Results: Latanoprost+BAK delivered by a drop induced cytotoxicity, cytoplasmic shrinkage, and loss of cell-cell contact, and expression of chemokine (C-C motif) ligand 2 and interleukin-6. In contrast, latanoprost+BAK delivered by the Optejet technology was both well tolerated and similar to no treatment controls and BAK-free latanoprost treatment. Conclusions: A microdose of latanoprost+BAK ophthalmical spray administered with the Optejet technology prevented the cytotoxicity associated with larger volumes found in eye drops. Precision dosing by the Optejet technology has the potential to decrease ocular surface disorder typically associated with eye drops containing preservatives.

Dendritic cell-derived lncRNAs in patients with acute coronary syndrome.

Long non-coding RNAs (lncRNAs) and dendritic cells (DC) play crucial roles in the development of acute coronary syndrome (ACS); however, the mechanisms remain unclear. To investigate this, we analysed the differentially expressed lncRNAs in monocyte-derived DCs (moDCs) from patients with ACS. Peripheral blood mononuclear cells were transformed into moDCs. Cellular morphology and expression levels of moDC-specific markers (CD80, CD86, CD11c, CD14 and HLA-DR) were analysed using electron microscopy (EM) and flow cytometry (FCM), respectively. Differentially expressed lncRNAs and their functions were predicted using gene sequencing, gene ontology and the Kyoto Encyclopedia of Genes and Genomes. The expression levels of markers, signalling pathway molecules (p-PI3K and p-AKT), inflammatory cytokines (IL-6 and IL-12p70) and target gene (C-C motif chemokine ligand (CCL) 15 and CCL14) were analysed by overexpression or silencing of candidate lncRNAs. EM revealed the cells to be suspended in dendritic pseudopodia. CD11c and HLA-DR were upregulated, while CD80 and CD86 were downregulated. Comparison between the UA versus ST group showed the highest number of differentially expressed lncRNAs (n = 113), followed by UA versus NST (n = 115), CON versus NST (n = 49) and CON versus ST (n = 35); however, the number was low for CON versus UA and ST versus NST groups. moDC-specific marker expression, signalling pathway molecules, inflammatory cytokines and CCL14 were upregulated following lentiviral overexpression of smart silencer-CCL15-CCL14; however, expression levels decreased following transfection with siRNA. The morphology, function and lncRNA expression of moDCs differ depending on the type of ACS. The differentially expressed lncRNAs, particularly CCL15-CCL14, regulate the function of moDCs. Thus, our study provides new insights regarding the role of lncRNAs in ACS and indicates the potential use of CCL15-CCL14 as a novel diagnostic marker and therapeutic target.