Axonal degeneration is an active process that has been associated with neurodegenerative conditions triggered by mechanical, metabolic, infectious, toxic, hereditary and inflammatory stimuli. This ...degenerative process can cause permanent loss of function, so it represents a focus for neuroprotective strategies. Several signaling pathways are implicated in axonal degeneration, but identification of an integrative mechanism for this self-destructive process has remained elusive. Here, we show that rapid axonal degeneration triggered by distinct mechanical and toxic insults is dependent on the activation of the mitochondrial permeability transition pore (mPTP). Both pharmacological and genetic targeting of cyclophilin D, a functional component of the mPTP, protects severed axons and vincristine-treated neurons from axonal degeneration in ex vivo and in vitro mouse and rat model systems. These effects were observed in axons from both the peripheral and central nervous system. Our results suggest that the mPTP is a key effector of axonal degeneration, upon which several independent signaling pathways converge. Since axonal and synapse degeneration are increasingly considered early pathological events in neurodegeneration, our work identifies a potential target for therapeutic intervention in a wide variety of conditions that lead to loss of axons and subsequent functional impairment.
To assess the effect of hydroxypropyl (HP)-Guar added to regular post-phacoemulsification treatment in dry eye signs and symptoms, and its influence on the expression of various inflammatory markers ...by flow cytometry (FCM) in impression cytology specimens.
This prospective, interventional, single-centre study included 48 eyes of 48 patients with age-related cataract. After phacoemulsification, patients were randomised to the usual treatment group (UT), with 21 patients who received tobramycin and dexamethasone eye drops (Tobradex, Alcon Cusí, Spain), and the HP-Guar group, with 27 patients who received the UT plus preservative-free artificial tears (Systane UD, Alcon Cusí, Spain). Corneal and conjunctival staining with fluorescein and lissamine green, tear film break-up time (TBUT), Schirmer's I test with anaesthesia (Jones test), tear clearance, and ocular surface disease index (OSDI) were assessed preoperatively and 1 month after surgery. Besides, conjunctival impression cytology was performed in order to investigate inflammatory markers (CD3, CD11b, and HLA-DR) using FCM.
HP-Guar group shows statistical better results compared with the UT group in TBUT (6.4+/-0.7 vs 9+/-2.5, P=0.0004), OSDI (11.5+/-8.2 vs 3.3+/-2.5, P=0.0002), ocular symptoms subscale (7.3+/-6.1 vs 1.7+/-1.8, P=0.0004), vision-related function subscale (2.2+/-1.8 vs 0.4+/-0.6, P=0.0002), CD3 (2.5+/-1.4 vs 1.1+/-1.1, P=0.011), and HLA-DR (6.8+/-4.5 vs 1.8+/-1.7, P=0.0002).
The addition of HP-Guar to regular treatment after cataract surgery reduces ocular surface inflammation and dry eye signs and symptoms.
Maintenance of the mitochondrial membrane potential (Deltapsim) is fundamental for the normal performance and survival of cells such as cardiomyocytes, that have a high energy requirement. ...Measurement of Deltapsim is therefore essential in order to develop an understanding of the molecular mechanisms controlling cardiomyocyte function. Here we have evaluated various potentiometric dyes for their ability to detect alterations of Deltapsim, using flow cytometry and confocal microscopy.
Primary cultures of cardiomyocytes from neonate rats were treated with mitochondrial uncouplers before or after loading with Rho123, DiOC(6)(3), CMXRos or JC-1, and then analysed by flow cytometry. Apoptotic cells were identified by light scatter and Annexin V staining.
The four potentiometric dyes tested were able to discriminate between viable and apoptotic cells. However, only JC-1 was able to detect the collapse of Deltapsim induced by uncouplers of mitochondrial respiration. Confocal microscopic analysis confirmed that JC-1 stained mitochondria in a potential-dependent manner. In contrast, CMXRos stained cardiomyocytes irrespective of alterations in Deltapsim.
We conclude that JC-1 is the optimal dye to use when measuring Deltapsim in cardiomyocytes.
An increasing number of reports indicate that single-celled organisms are able to die following what seems to be an ordered program of cell death with strong similarities to apoptosis from higher ...eukaryotes. DNA degradation and several other apoptotic-like processes have also been described in the parasitic protozoa Leishmania. However, the existence of an apoptotic death in this parasite is still a matter of controversy. Our results indicate that most of the processes of macromolecular degradation and organelle dysfunction observed in mammalian cells during apoptosis can also be reproduced in promastigotes of the genus Leishmania when incubated at temperatures above 38°C. These processes can be partially reversed by the expression of the anti-apoptotic mammalian gene Bcl-X^sub L^, which suggests that this family of apoptosis-regulating proteins was present very early in the evolution of eukaryotic cells.PUBLICATION ABSTRACT
Abstract
Study question
Is the miRNAs cargo of the extracellular vesicles (EVs) present in seminal plasma different between healthy semen donors and infertile patients?
Summary answer
There are ...significant differences in the miRNAs expression patterns in the load of seminal EVs between infertile patients and healthy semen donors
What is known already
Extracellular vesicles (EVs) present in seminal plasma contain a wide range of different proteins, lipids and nucleic acids, such as microRNA. These microRNA can execute their action by regulating gene expression of target cells triggering an important role in different reproductive processes. It is well stablished in different pathologies that the EVs cargo is representative for the functional status of the producer cell. In this sense, the alteration in microRNAs expression patterns in seminal plasma has been associated with spermatogenic impairments, subfertility and azoospermia, indicating that microRNAs in seminal EVs could work as biomarkers of the male fertility status
Study design, size, duration
This prospective study, performed between March 2021 to December 2022, included 30 semen samples from infertile males seeking infertility treatment (study group), and 7 semen samples from healthy semen donors (control group) from our donor semen bank. All the semen samples from patients showed abnormal seminal parameters (oligo- and/or asthenozoospermia). Men under drug treatment and/or with infectious or chronic diseases were excluded from the study
Participants/materials, setting, methods
After conventional semen analysis, EVs were isolated from seminal plasma by ultracentrifugation. To confirm EVs isolation, an aliquot of each sample was used for nanoparticle tracking analysis (NTA). Then, microRNAs from EVs were extracted and miRNAseq was performed. Raw data were standardized using counts per millions and ANOVA was performed to assess differences in the miRNAs expression between groups.Target genes were annotated using miRTarBase and functional enrichment analysis was performed by MIENTURNET and GeneCards
Main results and the role of chance
The presence of EVs after ultracentrifugation of seminal plasma was confirmed by NTA, showing a range size within 40-140 nm. miRNAseq generated reads for 914 annotated miRNAs and significant differences in the expression of 17 of them were found between patient and control samples. These miRNAs target a total of 1310 genes and functional enrichment analysis indicated that they participate in several processes related to cell differentiation, cell proliferation, immunomodulation, cellular adhesion and apoptosis. Specifically, patient group showed significant higher expression of some members of the hsa-let-7 family (p < 0.001 for let-7i-5p, let-7f-5p, let-7c-5p and let-7a-5p; p < 0.05 for let-7b-5p), which are implicated in adaptative immune system activation and differentiation by gene modulation. Results also found significant (p < 0.01) higher expression in patient group of hsa-miR-92a-3p, hsa-miR-200c-3p and hsa-miR-888-5p, which target genes are related to apoptotic processes. Finally, patient group showed significant (p < 0.001) lower expression of hsa-miR-320b and miR-320a-3p, which interact with transcription factor sequences (DLX5, MYC) implicated in cell proliferation and differentiation; and also lower expression in hsa-miR-423-5p (p < 0.01) and miR-423-3p (p < 0.05) that interact with the sequence of transcription factors as p21, a regulator of cell cycle progression at G1, and with RNA-binding proteins involved in ribosome assembly.
Limitations, reasons for caution
This study included males with dissimilar seminal disorders (astheno, oligo, and teratozoospermia). Therefore, further studies are needed lowering the heterogeneity of the study group. At present, no data about reproductive outcome is available
Wider implications of the findings
This work suggests that the significant differences found in the expression pattern in microRNAs in seminal EVs between infertile and fertile men may be related with gene modulation in the reproductive processes and could be used as novel biomarkers of male fertility status.
Trial registration number
1904-MAD-045-AP
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