bluuebunny/crossref_metadata_2025_split huggingface dataset:
Selective Stimulation of Inhibitory Motorneurons in the Mouse Colon Following Light Activation of Channel Rhodopsin‐2 (ChR2) Ex Vivo
Authors: Perez‐Medina Alberto , Manfredsson Fredric , Galligan James 
Published: November 2023

Inhibitory motor neurons in the enteric nervous system (ENS) are responsible for producing relaxations of gastrointestinal (GI) smooth muscle. Neurogenic relaxations are required to produce the pressure gradients needed to propel GI content along the gut. Nitric oxide (NO) and ATP (or related purine) are the predominant neurotransmitters released by inhibitory motorneurons in the ENS. Immunohistochemical studies using antibodies against nitric oxide synthase (NOS) and the vesicular nucleotide transporter (VNUT), protein markers for nitrergic and purinergic inhibitory motorneurons respectively, showed that there are NOS+ neurons, VNUT+ neurons and NOS/VNUT+ neurons in the myenteric plexus of the mouse colon. To determine if NO and ATP were released by the same or separate neurons, we injected an adeno‐associated virus (AAV9) containing a construct with genes encoding ChR2 and yellow fluorescent protein (YFP) bordered by loxP sites into the proximal colon of NOS‐cre expressing mice. 4–6 weeks later, the colon was harvested for electrophysiological measurements of circular muscle inhibitory junction potentials (IJPs, the electrophysiological correlate of muscle relaxation) evoked by blue light (470 nm) and transmural electrical stimulation. IJPs were recorded using intracellular microelectrodes. Light stimulation (5–20 mW/mm2) evoked biphasic IJPs in tissues from all mice successfully transduced by AAV9 (as measured by detection of the YFP reporter), but not in non‐transduced tissues (lacking YFP). Electrical stimulation evoked IJPs in tissues from all mice. The amplitude and duration of the light‐ and electrically‐evoked IJPs increased with the number of stimuli (1–7 stimuli). The fast, early phase of light‐ and electrically‐evoked IJPs was blocked by MRS2179 (10 μM, a P2Y1 receptor antagonist) while the slow, late phase of the IJP was blocked by nitro‐L‐arginine (100 μM, NOS inhibitor). IJPs were blocked completely by combined application of MRS2179 and nitro‐L‐arginine. These data provide direct evidence that inhibitory motorneurons in the mouse colon co‐release NO and ATP (or related purine) to cause relaxation of colon circular smooth muscle. Selective expression of ChR2 in subtypes of neurons will enable more precise identification of synaptic mechanisms and nerve pathways in the ENS.Support or Funding InformationSupported by R01DK094932
Neuroprotective Potential of Transient Receptor Potential Melastatin‐2channel Inhibitor Flufenamic Acid in Experimental Model of Focal Cerebral Ischemia
Authors: Medhi Bikash , Harikrishnareddy Dibbanti 
Published: November 2023

IntroductionCerebral Ischemia remains third leading cause of death and disability worldwide. Although progress has been made in supportive care, efforts to guard the brain from ischemic neuronal death have failed. Recently transient receptor potential melastatin‐2 (TRPM‐2) ion channel have been identified as major culprit that initiates neuronal death in cerebral stroke. These channels raises the intracellular Ca2+ levels and oxidative stress there by initiating mitochondrial membrane disruption, endoplasmic reticulum stress, glutamate excitotoxicity which initiates neuronal death. We therefore investigated the neuroprotective effect of TRPM‐2 channel inhibitor flufenamic acid (FA) for the treatment of Cerebral Ischemia.Materials and MethodsAdult male Wistar rats weighing 250±20g were divided into six different groups viz sham operated group, vehicle treated group, four different doses of flufenamic acid (FA‐5mg/kg, FA‐10mg/kg, FA‐20mg/kg, and FA‐40mg/kg groups). Focal Cerebral Ischemia was induced by middle cerebral artery occlusion (MCAO) model. Neurobehavioral assessmentwere monitored by Longa et al., five point scale with 10 grading scores, infarct size was calculated by using of 2,3,5‐triphenyltetrazolium chloride (TTC) staining, oxidative stress parameters were estimated by biochemical parameters, and brain water content was measured by wet‐dry method.ResultsFA showed neuroprotection in a dose dependent manner. FA 20mg significantly diminished neurological deficits and reduced infarct size in rats. Further, an early calcium dependent rise in levels of MDA was also found to be significantly reduced in ischemic brain regions following FA treatment. With higher doses of FA brain edema was also significantly reduced.ConclusionThe present study thus provide neuroprotective profile of FA in focal Cerebral Ischemia with its selective effect onTRPM‐2 channel.Support or Funding InformationPost Graduate Institute of Medical Education and Research
Subcellular localization and relevant mechanisms of human cancer‐related micropeptides
Authors: Deng Jing , Xu Wenli , Jie Yusheng , Chong Yutian 
Published: November 2023

AbstractRapid advances in high‐quality sequencing and bioinformatics have invalidated the argument that noncoding RNAs (ncRNAs) are junk transcripts that do not encode proteins. Increasing evidence suggests that small open reading frames (sORFs) in ncRNAs can encode micropeptides and polypeptides within 100 amino acids in length. Several micropeptides have been characterized and proven to have various functions in human physiology and pathology, particularly in cancer. The present review mainly highlights the latest studies on ncRNA‐encoded micropeptides in different cancers and categorizes them based on their subcellular localization, thereby providing a theoretical basis for micropeptide applications in the early diagnosis and prognosis of cancer and as therapeutic targets. However, considering the inherent characteristics of micropeptides and the limitations of the assay technology methods, more detailed information is warranted.
Acute, pro‐contractile effects of prorenin on rat mesenteric arteries
Authors: Rognant Salomé , Baldwin Samuel N. , Pritchard Harry A. T. , Greenstein Adam , Calloe Kirstine , Aalkjaer Christian , Jepps Thomas A. 
Published: November 2023

AbstractProrenin and the prorenin receptor ((P)RR) are important, yet controversial, members of the renin–angiotensin–aldosterone system. The ((P)RR) is expressed throughout the body, including the vasculature, however, the direct effect of prorenin on arterial contractility is yet to be determined. Within rat mesenteric arteries, immunostaining and proximity ligation assays were used to determine the interacting partners of (P)RR in freshly isolated vascular smooth muscle cells (VSMCs). Wire myography examined the functional effect of prorenin. Simultaneous changes in [Ca2+]i and force were recorded in arteries loaded with Fura‐2AM. Spontaneously transient outward currents were recorded via perforated whole‐cell patch‐clamp configuration in freshly isolated VSMCs. We found that the (P)RR is located within a distance of less than 40 nm from the V‐ATPase, caveolin‐1, ryanodine receptors, and large conductance Ca2+‐activated K+ channels (BKCa) in VSMCs. [Ca2+]i imaging and isometric tension recordings indicate that 1 nM prorenin enhanced α1‐adrenoreceptor‐mediated contraction, associated with an increased number of Ca2+ waves, independent of voltage‐gated Ca2+ channels activation. Incubation of VSMCs with 1 nM prorenin decreased the amplitude and frequency of spontaneously transient outward currents and attenuated BKCa‐mediated relaxation. Inhibition of the V‐ATPase with 100 nM bafilomycin prevented prorenin‐mediated inhibition of BKCa‐derived relaxation. Renin (1 nM) had no effect on BKCa‐mediated relaxation. In conclusion, prorenin enhances arterial contractility by inhibition of BKCa and increasing intracellular Ca2+ release. It is likely that this effect is mediated through a local shift in pH upon activation of the (P)RR and stimulation of the V‐ATPase.
The plow technique: An alternative method for the transection of liver parenchyma
Authors: Cotsoglou Christian , Bruno Federica , Lomaglio Laura , Cuttin Maria Serena , Bovo Giorgio , Gjoni Elson , Germini Alessandro , Paleino Sissi , Granieri Stefano 
Published: November 2023

Background:
Over the past 30 years, improvements in both technical skills and surgical equipment have been made to guarantee safe and effective liver parenchymal transection. In the present study, we propose a transection method, called “The Plow Technique” based on monopolar spray electrocoagulation in open hepatobiliary surgery.

Methods:
We conducted histological analyses on a cadaveric human liver measuring the diameters of all vascular structures at increasing depths of liver parenchyma. Furthermore, we retrospectively analyzed the data of 60 consecutive patients who underwent major and minor liver resections at our Institution.

Results:
The histological assessment of the distribution of vascular structures at different parenchymal depth points, failed to point out significant differences. Nevertheless, a trend toward an increased proportion of small caliber arterioles and centrilobular venules was found going from the Glissonian capsule to the deepest portion of liver parenchyma. All the hemorrhagic complications in our series were due to large caliber blood vessels spillage from the deeper parenchymal portions.

Conclusion:
The Plow technique may represent a feasible and safe technique for adequate coagulation and sealing of small vessels of the first 3 cm of the anterior surface of the liver. Besides its safety and effectiveness, it may help in speeding up parenchymal transection.
Survival benefit of liver resection following complete response to transarterial chemoembolization for intermediate-stage hepatocellular carcinoma: a retrospective, multicenter, cohort study
Authors: Hu Zili , Wang Xiaohui , Fu Yizhen , Yang Dinghua , Zhou Zhongguo , Chen Minshan , Song Xin , Zhang Yaojun 
Published: November 2023

Background:
High rate of tumor recurrence jeopardized the long-term survival of hepatocellular carcinoma (HCC) patients with complete response to transarterial chemoembolization (TACE). This study aims to evaluate the survival benefit of liver resection (LR) following the complete response to TACE for intermediate-stage HCC.

Methods:
A total of 281 intermediate-stage HCC patients with complete response to TACE followed by persistent observation (TACE group) or LR (TLR group) from January 01, 2011 to December 31, 2021 from three institutions in China were included. Overall survival (OS) and disease-free survival (DFS) of patients were compared between the two groups by propensity score–matching (PSM).

Results:
After PSM, the 1-, 3-, and 5-year OS rates were 91.4%, 71.5%, and 57.1% in the TACE group, and 96.6%, 81.8%, and 72.1% in the TLR group. The 1-, 3-, and 5-year DFS rates were 50.6%, 22.6%, and 6.8% in the TACE group, and 77.3%, 56.3%, and 38.7% in the TLR group. Compared with the TACE group, the TLR group showed significantly longer OS (HR, 0.528; 95%CI, 0.315–0.887; P=0.014) and DFS (HR, 0.388; 95%CI, 0.260–0.580; P<0.001). In patients beyond up-to-7 criterion, no difference was observed with OS (HR, 0.708; 95% CI, 0.354–1.419; P=0.329). LR following the complete response to TACE was safety.

Conclusions:
This study suggests that intermediate-stage HCC patients could benefit from LR following the complete response to TACE, resulting in longer OS and DFS. In addition, patients beyond up-to-7 could not benefit from the LR treatments.
Exploring the activity and the essentiality of the putative Δ6-desaturase in the procyclic and bloodstream forms ofTrypanosoma brucei
Authors: Cerone Michela , Smith Terry K 
Published: November 2023

AbstractTrypanosomatids have been shown to possess an exclusive and finely regulated biosynthetic pathway forde novosynthesis of fatty acids (FAs) and particularly of polyunsaturated fatty acids (PUFAs). The key enzymes for the process of unsaturation are known as desaturases. In this work, we explored the biocatalytic activity of the putative Δ6-desaturase (Tb11.v5.0580) in the native organismT. brucei. Utilising fatty acid analysisviaGC-MS, we were able to elucidateviagenetic manipulation of the level of expression of Δ6-desaturases in both procyclic (PCF) and bloodstream (BSF) forms ofT. bruceiandviasupplementation of the media with various levels of FA sources, that docosahexaenoic acid (22:6) and/or docosapentaenoic acid (22:5), and arachidonic acid (20:4) and/or docosatetraenoic acid (22:4) are the products and the substrates respectively of this Δ6-desaturases. Interestingly, we were able to observe,vialipidomic analysis with ESI-MS/MS, an increase in inositol-phosphoryl ceramide (IPC) in response to the overexpression of Δ6-desaturases in low-fat media, both in PCF and rather surprisingly in BSF. The formation of IPC is normally only observed in the stumpy and procyclic forms ofT. brucei. Therefore, the expression levels of Δ6-desaturases, which varies between BSF and PCF, might be involved in the cascade(s) of metabolic events that cause these remodelling of the lipid pools and ultimately morphological changes, which are key to the transition between these life-cycle stages.Author summaryTrypanosoma bruceiis a unicellular parasite that causes human and animal African trypanosomiasis. These parasites have the special ability to make their own pool of fat molecules by assembling and modifying the fatty acid building blocks that they take up from the human and animal hosts and from the insect vector. In this study, we investigated the unknown activity of a desaturase enzyme. By modulating its activity, we showed that it can make different levels of high-value long chain polyunsaturated fatty acids (LC-PUFAs) often known as omega-6 and omega-3. If we increase or reduce the fat sources available from the outer environment, the cells respond by making more or less LC-PUFAs and by forming different type of lipids and sphingolipids for their cellular membranes. We highlighted that by tuning the level of activity of the desaturase and varying the type and amounts of fat sources available to the cells,T. bruceican alter their morphology. This is key for the parasites to adapt to the various environments and the nutrients therein that are often constantly changing within the host, allowing the shift between different life-stages during the complex life cycle from the insect vector to the host and back.
The endogenousMtv8locus and the immunoglobulin repertoire
Authors: Beilinson Helen A. , Erickson Steven A. , Golovkina Tatyana 
Published: November 2023

AbstractThe vast diversity of mammalian adaptive antigen receptors allows for robust and efficient immune responses against a wide number of pathogens. The antigen receptor repertoire is built during the recombination and hypermutation of B and T cell receptor (BCR, TCR) loci. V(D)J recombination rearranges these antigen receptor loci, which are organized as an array of separate V, (D), and J gene segments. Transcription activation at the recombining locus leads to changes in the local three-dimensional architecture, which subsequently contributes to which gene segments are utilized for recombination. The endogenous retrovirus (ERV) mouse mammary tumor provirus 8 (Mtv8) resides on mouse chromosome 6 interposed within the large array of light chain kappa V gene segments. As ERVs contribute to changes in genomic architecture by driving high levels of transcription of neighboring genes, it was suggested thatMtv8could influence the BCR repertoire. We generatedMtv8-deficient mice to determine if the ERV influences V(D)J recombination to test this possibility. We find thatMtv8does not influence the BCR repertoire.
Canonical and noncanonical contribution of thyroid hormone receptor isoforms alpha and beta to cardiac hypertrophy and heart rate in male mice
Authors: Geist Daniela , Hönes G. Sebastian , Grund Susanne C. , Pape Janina , Siemes Devon , Spangenberg Philippa , Tolstik Elen , Dörr Stefanie , Spielmann Nadine , Fuchs Helmut , Gailus-Durner Valerie , Hrabe de Angelis Martin , Mittag Jens , Engel Daniel R. , Führer Dagmar , Lorenz Kristina , Moeller Lars C. 
Published: November 2023

AbstractBackgroundStimulation of ventricular hypertrophy and heart rate are two major cardiac effects of thyroid hormone (TH). Aim of this study was to determinein vivowhich TH receptor (TR), α or β, and which mode of TR action, canonical gene expression or DNA-binding independent noncanonical action, mediate these effects.Material and methodsWe compared global TRα and TRβ knockout mice (TRαKO; TRβKO) with WT mice to determine the TR isoform responsible for T3 effects. The relevance of TR DNA- binding was studied in mice with a mutation in the DNA-binding domain that selectively abrogates DNA binding and canonical TR action (TRαGS; TRβGS). Hearts were studied with echocardiography at baseline and after seven weeks T3-treatment. Gene expression was measured with real-time PCR. Heart rate was recorded with radiotelemetry transmitters for seven weeks in untreated, hypothyroid and T3-treated mice.ResultsT3 induced ventricular hypertrophy in WT and TRβKOmice, but not in TRαKOmice. Hypertrophy was also induced in TRαGSmice. Thus, hypertrophy is mostly mediated by noncanonical TRα action. Similarly, repression ofMhy7occurred in WT and TRαGSmice. Basal heart rate was largely dependent on canonical TRα action. But responsiveness to hypothyroidism and T3-treatment as well as expression of pacemaker geneHcn2were still preserved in TRαKOmice, demonstrating that TRβ could compensate for absence of TRα.ConclusionT3-induced cardiac hypertrophy could be attributed to noncanonical TRα action, whereas heart rate regulation was mediated by canonical TRα action. TRβ could substitute for canonical, but not noncanonical TRα action.
CryoET reveals actin filaments within platelet microtubules
Authors: Tsuji Chisato , Bradshaw Marston , Allen Megan , Jackson Molly L. , Mantell Judith , Borucu Ufuk , Poole Alastair W. , Verkade Paul , Hers Ingeborg , Paul Danielle M. , Dodding Mark P. 
Published: November 2023

AbstractCrosstalk between the actin and microtubule cytoskeletons is essential for many cellular processes. Recent studies have shown that microtubules and F-actin can assemble to form a composite structure where F-actin occupies the microtubule lumen. Whether these cytoskeletal hybrids exist in physiological settings and how they are formed is unclear. Here, we show that the short-crossover Class I actin filament previously identified inside microtubules in human HAP1 cells is cofilin-bound F-actin. Lumenal F-actin can be reconstitutedin vitro, but cofilin is not essential. Moreover, actin filaments with both cofilin-bound and canonical morphologies reside within human platelet microtubules under physiological conditions. We propose that stress placed upon the microtubule network during motor-driven microtubule looping and sliding may facilitate the incorporation of actin into microtubules.
TRPS1 maintains luminal progenitors in the mammary gland by repressing SRF/MRTF activity
Authors: Tollot-Wegner Marie , Jessen Marco , Kim KyungMok , Sanz-Moreno Adrián , Spielmann Nadine , Gailus-Durner Valerie , Fuchs Helmut , de Angelis Martin Hrabe , von Eyss Björn 
Published: November 2023

AbstractThe transcription factor TRPS1 is a context-dependent oncogene in breast cancer [1] [2] [3] [4] [5]. In the mammary gland, TRPS1 activity is restricted to the luminal population and is critical during puberty and pregnancy [2]. Its function in the resting state remains however unclear. To evaluate whether it could be a target for cancer therapy, we investigated TRPS1 function in the healthy adult mammary gland using a conditional ubiquitous depletion mouse model where long-term depletion does not affect fitness. We show that TRPS1 activity is essential to maintain a functional luminal progenitor compartment. This requires the repression of both YAP/TAZ and SRF/MRTF activities, TRPS1 represses SRF/MRTF activity indirectly by modulating RhoA activity. Our work uncovers a hitherto undisclosed function of TRPS1 in luminal progenitors intrinsically linked to mechanotransduction in the mammary gland. It also provides new insights into the oncogenic functions of TRPS1 as luminal progenitors are likely the cells of origin of many breast cancers.Significance statementThe transcription factor TRPS1 is a context-dependent oncogene in breast cancer. It is unclear how TRPS1 contributes to cancer development and whether it could be a target for therapy. Here we established a mouse model mimicking the systemic effect of TRPS1 drug targeting. With this model, we can show that TRPS1 depletion does not impact the fitness of the animals and that the role of TRPS1 is to maintain a functional luminal progenitor pool in the mammary gland. Mechanistically, TRPS1 represses a mechano-transduction program preventing their commitment to an alveolar fate. Because there is growing evidence that breast cancer originates from the expansion of altered luminal progenitors, our work provides valuable insights into the understanding of breast cancer initiation.
Chaperone-mediated autophagy is an overlooked pathway for mutant α1-antitrypsin Z degradation
Authors: Lin Jiayu , Lu Haorui , Wei Xinyue , Dai Yan , Wu Rihan , Yang Hao , Rao Lang 
Published: November 2023

AbstractChaperone-mediated autophagy (CMA) is a specific form of autophagy that selectively targets proteins containing a KFERQ-like motif and relies on the chaperone protein HSC70 for substrate recognition. In α1-antitrypsin deficiency (AATD), a disease characterized by the hepatic build-up of α1-Antitrypsin Z mutant (ATZ), CMA’s role had been unclear. This work demonstrates the critical role that CMA plays in preventing ATZ accumulation; suppressing CMA worsens ATZ accumulation, whilst activating it through chemical stimulation or LAMP2A overexpression promotes ATZ breakdown. Specifically, ATZ’s 121QELLR125 motif is critical for HSC70 recognition and LAMP2A’s charged C-terminal cytoplasmic tail is vital for substrate binding, facilitating CMA-mediated degradation of ATZ. This selective activation of CMA operates independently from other autophagy pathways and alleviate ATZ aggregates caused cellular stress. These findings highlight CMA’s critical function in cellular protein quality control of ATZ and place it as a novel target for AATD treatment approaches.
Efficient scar-free knock-ins of several kilobases by engineered CRISPR/Cas endonucleases
Authors: Schreiber Tom , Prange Anja , Schäfer Petra , Iwen Thomas , Grützner Ramona , Marillonnet Sylvestre , Tissier Alain 
Published: November 2023

AbstractIn plants and mammals, non-homologous end-joining is the dominant pathway to repair DNA double strand breaks, making it challenging to generate knock-in events. Using a transient assay inNicotiana benthamianawe identified two groups of exonucleases, respectively from the Herpes Virus and from the bacteriophage T7 families, hat confer up to 38-fold increase in HDR frequencies when fused to Cas9. We achieved precise and scar-free insertion of several kilobases of DNA both in transient and stable transformation systems. InArabidopsis thaliana, fusion of Cas9 to a Herpes Virus family exonuclease leads to 10-fold higher frequencies of knock-ins. Our results open perspectives for the routine production of knock-in and gene replacement events in plants.One-Sentence SummaryFusions of CRISPR endonucleases to specific 5′-exonucleases leads to significant increase in scar-free multikilobase knock-ins.