Annual rhythm in immune functions of blood leucocytes in an ophidian, Natrix piscator.

Annual variations in animal's physiological functions are an essential strategy to deal with seasonal challenges which also vary according to the time of year. Information regarding annual adaptations in the immune-competence to cope with seasonal stressors in reptiles is scarce. The present research plan was designed to analyze the presence of circannual immune rhythms in defense responses of the leucocytes in an ophidian, Natrix piscator. Peripheral blood leucocytes were obtained, counted, and superoxide anion production, neutrophil phagocytosis, and nitrite release were tested to assess the innate immune functions. Peripheral blood lymphocytes were separated by centrifugation (utilizing density gradient) and the cell proliferation was measured. The Cosinor rhythmometry disclosed the presence of significant annual rhythms in the number of leucocytes, superoxide anion production, nitric oxide production, and proliferation of stimulated lymphocytes. The authors found that respiratory burst activity and proliferative responses of lymphocytes were crucial immune responses that showed the annual rhythm. It was summarized that the immune function of the N. piscator is a labile attribute that makes the animal competent to cope with the seasonal stressor by adjustment in the potency of response.

A Systems Biology Approach Unveils a Critical Role of DPP4 in Upper Gastrointestinal Cancer Patient Outcomes.

Gastrointestinal (GI) cancers comprise of cancers that affect the digestive system and its accessory organs. The late detection and poor prognosis of GI cancer emphasizes the importance of identifying reliable and precise biomarkers for early diagnosis and prediction of prognosis. The membrane-bound glycoprotein dipeptidyl-peptidase 4 (DPP4), also known as CD26, is ubiquitously expressed and has a wide spectrum of biological roles. The role of DPP4/CD26 in tumor progression in different types of cancers remains elusive. However, the link between DPP4 and tumor-infiltrating cells, as well as its prognostic significance in malignancies, still require further investigation. This study was intended to elucidate the correlation of DPP4 expression and survival along with prognosis, followed by its associated enriched molecular pathways and immune cell marker levels in upper GI cancers. Results demonstrated a strong correlation between increased DPP4 expression and a worse prognosis in esophageal and gastric cancer and the co-expressed common genes with DPP4 were associated with crucial molecular pathways involved in tumorigenesis. Additionally, DPP4 was shown to be significantly linked to several immune infiltrating cell marker genes, including Macrophages (M1, M2 and Tumor Associated Macrophages), neutrophils, Treg, T-cell exhaustion, Th1 and Th2. Overall, our findings suggest that DPP4 may serve as a substantial prognostic biomarker, a possible therapeutic target, as well as it can play a critical role in the regulation of immune cell invasion in patients with gastroesophageal (esophageal, gastroesophageal junction and gastric) cancer. KEY WORDS: DPP4, integrated analysis, GI cancer, gastroesophageal cancer, gastroesophageal junction, prognosis.

Recurring SARS-CoV-2 variants: an update on post-pandemic, co-infections and immune response.

The post-pandemic era following the global spread of the SARS-CoV-2 virus has brought about persistent concerns regarding recurring coinfections. While significant strides in genome mapping, diagnostics, and vaccine development have controlled the pandemic and reduced fatalities, ongoing virus mutations necessitate a deeper exploration of the interplay between SARS-CoV-2 mutations and the host's immune response. Various vaccines, including RNA-based ones like Pfizer and Moderna, viral vector vaccines like Johnson & Johnson and AstraZeneca, and protein subunit vaccines like Novavax, have played critical roles in mitigating the impact of COVID-19. Understanding their strengths and limitations is crucial for tailoring future vaccines to specific variants and individual needs. The intricate relationship between SARS-CoV-2 mutations and the immune response remains a focus of intense research, providing insights into personalized treatment strategies and long-term effects like long-COVID. This article offers an overview of the post-pandemic landscape, highlighting emerging variants, summarizing vaccine platforms, and delving into immunological responses and the phenomenon of long-COVID. By presenting clinical findings, it aims to contribute to the ongoing understanding of COVID-19's progression in the aftermath of the pandemic.

Long Non-Coding RNAs: New Insights in Neurodegenerative Diseases.

Neurodegenerative diseases (NDDs), including Alzheimer's disease (AD), Parkinson's disease (PD), and amyotrophic lateral sclerosis (ALS), are gradually becoming a burden to society. The adverse effects and mortality/morbidity rates associated with these NDDs are a cause of many healthcare concerns. The pathologic alterations of NDDs are related to mitochondrial dysfunction, oxidative stress, and inflammation, which further stimulate the progression of NDDs. Recently, long non-coding RNAs (lncRNAs) have attracted ample attention as critical mediators in the pathology of NDDs. However, there is a significant gap in understanding the biological function, molecular mechanisms, and potential importance of lncRNAs in NDDs. This review documents the current research on lncRNAs and their implications in NDDs. We further summarize the potential implication of lncRNAs to serve as novel therapeutic targets and biomarkers for patients with NDDs.

Renal Mitochondrial ATP Transporter Ablation Ameliorates Obesity-Induced CKD.

SIGNIFICANCE STATEMENT: This study sheds light on the central role of adenine nucleotide translocase 2 (ANT2) in the pathogenesis of obesity-induced CKD. Our data demonstrate that ANT2 depletion in renal proximal tubule cells (RPTCs) leads to a shift in their primary metabolic program from fatty acid oxidation to aerobic glycolysis, resulting in mitochondrial protection, cellular survival, and preservation of renal function. These findings provide new insights into the underlying mechanisms of obesity-induced CKD and have the potential to be translated toward the development of targeted therapeutic strategies for this debilitating condition. BACKGROUND: The impairment in ATP production and transport in RPTCs has been linked to the pathogenesis of obesity-induced CKD. This condition is characterized by kidney dysfunction, inflammation, lipotoxicity, and fibrosis. In this study, we investigated the role of ANT2, which serves as the primary regulator of cellular ATP content in RPTCs, in the development of obesity-induced CKD. METHODS: We generated RPTC-specific ANT2 knockout ( RPTC-ANT2-/- ) mice, which were then subjected to a 24-week high-fat diet-feeding regimen. We conducted comprehensive assessment of renal morphology, function, and metabolic alterations of these mice. In addition, we used large-scale transcriptomics, proteomics, and metabolomics analyses to gain insights into the role of ANT2 in regulating mitochondrial function, RPTC physiology, and overall renal health. RESULTS: Our findings revealed that obese RPTC-ANT2-/- mice displayed preserved renal morphology and function, along with a notable absence of kidney lipotoxicity and fibrosis. The depletion of Ant2 in RPTCs led to a fundamental rewiring of their primary metabolic program. Specifically, these cells shifted from oxidizing fatty acids as their primary energy source to favoring aerobic glycolysis, a phenomenon mediated by the testis-selective Ant4. CONCLUSIONS: We propose a significant role for RPTC-Ant2 in the development of obesity-induced CKD. The nullification of RPTC-Ant2 triggers a cascade of cellular mechanisms, including mitochondrial protection, enhanced RPTC survival, and ultimately the preservation of kidney function. These findings shed new light on the complex metabolic pathways contributing to CKD development and suggest potential therapeutic targets for this condition.

Time Kills: Impact of Socioeconomic Deprivation on Timely Access to Guideline-Concordant Treatment in Foregut Cancer.

BACKGROUND: Receipt of guideline-concordant treatment (GCT) is associated with improved prognosis in foregut cancers. Studies show that patients living in areas of high neighborhood deprivation have worse healthcare outcomes; however, its effect on GCT in foregut cancers has not been evaluated. We studied the impact of the area deprivation index (ADI) as a barrier to GCT. STUDY DESIGN: A single-institution retrospective review of 498 foregut cancer patients (gastric, pancreatic, and hepatobiliary adenocarcinoma) from 2018 to 2022 was performed. GCT was defined based on National Comprehensive Cancer Network guidelines. ADI, a validated measure of neighborhood disadvantage was divided into terciles (low, medium, and high) with high ADI indicating the most disadvantage. RESULTS: Of 498 patients, 328 (66%) received GCT: 66%, 72%, and 59% in pancreatic, gastric, and hepatobiliary cancers, respectively. Median (interquartile range) time from symptoms to workup was 6 (3 to 13) weeks, from diagnosis to oncology appointment was 4 (1 to 10) weeks, and from oncology appointment to treatment was 4 (2 to 10) weeks. Forty-six percent were diagnosed in the emergency department. On multivariable analyses, age 75 years or older (odds ratio [OR] 0.39 [95% CI 0.18 to 0.87]), Black race (OR 0.52 [95% CI 0.31 to 0.86]), high ADI (OR 0.25 (95% CI 0.14 to 0.48]), 6 weeks or more from symptoms to workup (OR 0.44 [95% CI 0.27 to 0.73]), 4 weeks or more from diagnosis to oncology appointment (OR 0.76 [95% CI 0.46 to 0.93]), and 4 weeks or more from oncology appointment to treatment (OR 0.63 [95% CI 0.36 to 0.98]) were independently associated with nonreceipt of GCT. CONCLUSIONS: Residence in an area of high deprivation predicts nonreceipt of GCT. This is due to multiple individual- and system-level barriers. Identifying these barriers and developing effective interventions, including community outreach and collaboration, leveraging telehealth, and increasing oncologic expertise in underserved areas, may improve access to GCT.

Mutations associated with autism lead to similar synaptic and behavioral alterations in both sexes of male and female mouse brain.

Autism spectrum disorder (ASD) is a neurodevelopmental disorder based on synaptic abnormalities. The estimated prevalence rate of male individuals diagnosed with ASD prevails over females is in a proportion of 4:1. Consequently, males remain the main focus in ASD studies in clinical and experimental settings. Meanwhile, some studies point to an underestimation of this disorder in females. In this work, we studied the sex differences of the synaptic and behavioral phenotypes of ASD mouse models. Juvenile male and female Shank3Δ4-22 and Cntnap2-/- mutant mice and their WT littermates were used in the experiments. The animals were subjected to a Three-Chamber Sociability Test, then euthanized, and the whole cortex was used for the evaluation of the synaptic phenotype. Protein levels of glutamatergic (NR1) and GABAergic (GAD1 and VGAT) neuronal markers were measured. Protein level of synaptophysin (Syp) was also measured. Dendritic spine density in somatosensory neurons was analyzed by Golgi staining methods. Spine Density and GAD1, NR1, VGAT, and Syp levels were significantly reduced in Shank3Δ4-22 and Cntnap2-/- mice compared to the control group irrespective of sex, indicating impaired synaptic development in the mutant mice. These results were consistent with the lack of differences in the three-chamber sociability test between male and female mice. In conclusion, female ASD mice of both mutations undergo similar synaptic aberrations as their male counterparts and need to be studied along with the male animals. Finally, this work urges the psychiatry scientific community to use both sexes in their investigations.

Antimalarial potential, LC-MS secondary metabolite profiling and computational studies of Zingiber officinale.

Malaria is among the top-ranked parasitic diseases that pose a threat to the existence of the human race. This study evaluated the antimalarial effect of the rhizome of Zingiber officinale in infected mice, performed secondary metabolite profiling and detailed computational antimalarial evaluation through molecular docking, molecular dynamics (MD) simulation and density functional theory methods. The antimalarial potential of Z. officinale was performed using the in vivo chemosuppressive model; secondary metabolite profiling was carried out using liquid chromatography-mass spectrometry (LC-MS). Molecular docking was performed with Autodock Vina while the MD simulation was performed with Schrodinger desmond suite for 100 ns and DFT calculations with B3LYP (6-31G) basis set. The extract showed 64% parasitaemia suppression, with a dose-dependent increase in activity up to 200 mg/kg. The chemical profiling of the extract tentatively identified eight phytochemicals. The molecular docking studies with plasmepsin II and Plasmodium falciparum dihydrofolate reductase-thymidylate synthase (PfDHFR-TS) identified gingerenone A as the hit molecule, and MMGBSA values corroborate the binding energies obtained. The electronic parameters of gingerenone A revealed its significant antimalarial potential. The antimalarial activity elicited by the extract of Z. officinale and the bioactive chemical constituent supports its usage in ethnomedicine.Communicated by Ramaswamy H. Sarma.

Plasmepsin II inhibitory potential of phytochemicals isolated from African antimalarial plants: a computational approach.

Plamepsin II has been identified as a therapeutic target in the Plasmodium falciparum's life cycle and may lead to a drastic reduction in deaths caused by malaria worldwide. Africa flora is rich in medicinal qualities and possesses both simple and complex bioactive phytochemicals. This study utilized computational approaches like molecular docking, molecular dynamics simulation, quantum chemical calculations and ADMET to evaluate the plasmepsin II inhibitory properties of phytochemicals isolated from African antimalarial plants. Molecular docking was carried out to estimate the binding affinity of 229 phytochemicals whereby ekeberin A, dichamanetin, 10-hydroxyusambaresine, chamuvaritin and diuvaretin were selected. Further, RMSD and RMSF plots from the 100 ns simulation results showed that the screened phytochemicals were stable in the enzyme's binding pocket. The quantum chemical calculation revealed that all the phytochemicals are strong electrophiles, while ekeberin A was identified as the most stable and dichamanetin as the most reactive. Also, ADMET studies established the drug candidacy of the phytochemicals. Thus, these phytochemicals could act as good antimalarial agents after extensive in vitro and in vivo studies.Communicated by Ramaswamy H. Sarma.

MUC13 drives cancer aggressiveness and metastasis through the YAP1-dependent pathway.

Anchorage-independent survival after intravasation of cancer cells from the primary tumor site represents a critical step in metastasis. Here, we reveal new insights into how MUC13-mediated anoikis resistance, coupled with survival of colorectal tumor cells, leads to distant metastasis. We found that MUC13 targets a potent transcriptional coactivator, YAP1, and drives its nuclear translocation via forming a novel survival complex, which in turn augments the levels of pro-survival and metastasis-associated genes. High expression of MUC13 is correlated well with extensive macrometastasis of colon cancer cells with elevated nuclear YAP1 in physiologically relevant whole animal model systems. Interestingly, a positive correlation of MUC13 and YAP1 expression was observed in human colorectal cancer tissues. In brief, the results presented here broaden the significance of MCU13 in cancer metastasis via targeting YAP1 for the first time and provide new avenues for developing novel strategies for targeting cancer metastasis.

SCAPS study on the effect of various hole transport layer on highly efficient 31.86% eco-friendly CZTS based solar cell.

Copper Zinc Tin Sulphide (CZTS) is a propitious semiconductor for active absorber material in thin-film solar cells (SCs). Here, SC architecture comprising FTO/ZnS/CZTS/variable HTLs/Au is discussed. Fluorine-doped tin oxide (FTO) and gold (Au) are used as front and back contacts, respectively. Zinc sulphide (ZnS) is used as an active electron transport layer (ETL), while different Cu-based materials (Cu2O, CuO, CuI, and CuSCN) are used as hole transport layers (HTL). A one-dimensional solar cell capacitance simulator (SCAPS-1D) is utilized to simulate the SC structure. Among different Cu-based HTLs, Cu2O is preferred as a potential candidate for high cell performance of CZTS-based SC. The effects of various layer parameters such as thickness, doping density, and carrier concentrations, electron affinity of HTL and absorber, respectively, are also discussed. After optimization of the device, variation of operating temperature and the effect of series and shunt resistance are also taken into consideration. The optimized results of thickness and acceptor concentration (NA) of absorber material are 1.5 µm and approx. 1.0 × 1019 cm-3, respectively. In addition, the function of HTL (with and without) in the designed SC structure is also studied. Capacitance-voltage (C-V) characteristics are also discussed to get an insight of built-in potential. We have achieved cell performances viz. efficiency = 31.86%, short circuit current density = 32.05 mA/cm2, open circuit voltage = 1.19 V, and fill factor = 83.37%.

Nitric Oxide Synthase Inhibition Prevents Cell Proliferation in Glioblastoma.

Glioblastoma multiforme (GBM) is a prevalent and aggressive primary brain tumor, presenting substantial treatment challenges and high relapse rates. GBM is characterized by alterations in molecular signaling and enzyme expression within malignant cells. This tumor exhibits elevated nitric oxide (NO.) levels. NO. is a crucial signaling molecule involved in the regulation of neuronal functions, synaptic transmission, and cell proliferation. It is primarily synthesized from L-arginine by nitric oxide synthase (NOS) enzymes. The increased levels of NO. in GBM stem from dysregulated activity and expression of clinically relevant NOS isoforms, particularly inducible NOS (iNOS) and neuronal NOS (nNOS). Based on this knowledge, we hypothesize that targeted pharmacological intervention with N6-(1-iminoethyl)-L-lysine (L-NIL), an iNOS inhibitor, and 7-Nitroindazole (7-NI), an nNOS inhibitor, may suggest a promising therapeutic strategy for the treatment of GBM. To test our hypothesis, we utilized the U87-MG cell line as an in vitro model of GBM. Our results showed that treatment with L-NIL and 7-NI led to a reduction in NO. levels, NOS activity, and clonogenic proliferation in U87-MG cells. These findings suggest that NO. and NOS enzymes might be prospective therapeutic targets for GBM.

Effects of extended-release 7-nitroindazole gel formulation treatment on the behavior of Shank3 mouse model of autism.

Autism spectrum disorder (ASD) is a neurodevelopmental disorder characterized by behavioral deficits such as abnormalities in communication, social interaction, anxiety, and repetitive behavior. We have recently shown that the Shank3 mutation in mice representing a model of ASD causes excessive nitric oxide (NO) levels and aberrant protein S-nitrosylation. Further, 10-day daily injections of 7-NI, a neuronal nitric oxide synthase inhibitor, into Shank3Δ4-22 and Cntnap2(-/-) mutant mice (models of ASD) at a dose of 80 mg/kg reversed the manifestations of ASD phenotype. In this study, we proposed an extended release of 7-NI using a novel drug system. Importantly, unlike the intraperitoneal injections, our new preparation of poly (sebacic acid-co-ricinoleic acid) (PSARA) gel containing 7-NI was injected subcutaneously into the mutant mice only once. The animals underwent behavioral testing starting from day 3 post-injection. It should be noted that the developed PSARA gel formulation allowed a slow release of 7-NI maintaining the plasma level of the drug at ∼45 µg/ml/day. Further, we observed improved memory and social interaction and reduced anxiety-like behavior in Shank3 mutant mice. This was accompanied by a reduction in 3-nitrotyrosine levels (an indicator of nitrative/nitrosative stress) in plasma. Overall, we suggest that our single-dose formulation of PSARA gel is very efficient in rendering a therapeutic effect of 7-NI for at least 10 days. This approach may provide in the future a rational design of an effective ASD treatment using 7-NI and its clinical translation.

Seasonal Immune Rhythms of head kidney and spleen cells in the freshwater Teleost, Channa punctatus.

Annual rhythms in immune function are the reflection of a crucial physiological strategy to deal with environmental stressors. The fish are pivotal animal models to study the annual rhythm and to understand the evolution of the vertebrate biological system. The current research was planned to assess the annual changes in the innate immune functions of immune cells in a teleost, Channa punctatus. Head kidney and splenic macrophage phagocytosis, superoxide generation, and nitrite release were evaluated to assess innate immunity. Cell-mediated immunity was measured through head kidney and splenic lymphocyte proliferation in presence of mitogens. The superoxide anion generation by the cells of head kidney and spleen was maximum in October. A bimodal pattern in nitrite production was observed with the first peak in November and the second in March. Cosinor analysis revealed a statistically significant annual rhythm in nitrite production. Similarly, phagocytosis and lymphocyte proliferation also showed statistically significant annual rhythms. It was concluded that animals maintain an optimum immune response in seasonally changing environments. Elevated immunity during certain times of the year might assist animals deal with seasonal environmental stressors. Further research may be focused upon measuring survival rate and reproductive success after season induced elevated immunity.

A novel approach for rapid and sensitive detection of Zika virus utilizing silver nanoislands as SERS platform.

To control the spread of the disease, the Zika virus (ZIKV), a flavivirus infection spread by mosquitoes and common in across the world, needs to be accurately and promptly diagnosed. This endeavour gets challenging when early-stage illnesses have low viral loads. As a result, we have created a biosensor based on surface-enhanced Raman scattering (SERS) for the quick, accurate, and timely diagnosis of the Zika virus. In this study, a glass coverslip was coated with silver nanoislands, which were then utilized as the surface for creating the sensing platform. Silver nanoislands exhibit strong plasmonic activity and good conductive characteristics. It enhances the Raman signals as a result and gives the SERS platform an appropriate surface. The created platform has been applied to Zika virus detection. With a limit of detection (LOD) of 0.11 ng/mL, the constructed sensor exhibits a linear range from 5 ng/mL to 1000 ng/mL. Hence, even at the nanogram scale, this technique may be a major improvement over clinical diagnosis approaches for making proper, precise, and accurate Zika virus detection.

The NO Answer for Autism Spectrum Disorder.

Autism spectrum disorders (ASDs) include a wide range of neurodevelopmental disorders. Several reports showed that mutations in different high-risk ASD genes lead to ASD. However, the underlying molecular mechanisms have not been deciphered. Recently, they reported a dramatic increase in nitric oxide (NO) levels in ASD mouse models. Here, they conducted a multidisciplinary study to investigate the role of NO in ASD. High levels of nitrosative stress biomarkers are found in both the Shank3 and Cntnap2 ASD mouse models. Pharmacological intervention with a neuronal NO synthase (nNOS) inhibitor in both models led to a reversal of the molecular, synaptic, and behavioral ASD-associated phenotypes. Importantly, treating iPSC-derived cortical neurons from patients with SHANK3 mutation with the nNOS inhibitor showed similar therapeutic effects. Clinically, they found a significant increase in nitrosative stress biomarkers in the plasma of low-functioning ASD patients. Bioinformatics of the SNO-proteome revealed that the complement system is enriched in ASD. This novel work reveals, for the first time, that NO plays a significant role in ASD. Their important findings will open novel directions to examine NO in diverse mutations on the spectrum as well as in other neurodevelopmental disorders. Finally, it suggests a novel strategy for effectively treating ASD.

Machine-learning technique, QSAR and molecular dynamics for hERG-drug interactions.

One of the most well-known anti-targets defining medication cardiotoxicity is the voltage-dependent hERG K + channel, which is well-known for its crucial involvement in cardiac action potential repolarization. Torsades de Pointes, QT prolongation, and sudden death are all caused by hERG (the human Ether-à-go-go-Related Gene) inhibition. There is great interest in creating predictive computational (in silico) tools to identify and weed out potential hERG blockers early in the drug discovery process because testing for hERG liability and the traditional experimental screening are complicated, expensive and time-consuming. This study used 2D descriptors of a large curated dataset of 6766 compounds and machine learning approaches to build robust descriptor-based QSAR and predictive classification models for KCNH2 liability. Decision Tree, Random Forest, Logistic Regression, Ada Boosting, kNN, SVM, Naïve Bayes, neural network and stochastic gradient classification classifier algorithms were used to build classification models. If a compound's IC50 value was between 10 µM and less, it was classified as a blocker (hERG-positive), and if it was more, it was classified as a non-blocker (hERG-negative). Matthew's correlation coefficient formula and F1score were applied to compare and track the developed models' performance. Molecular docking and dynamics studies were performed to understand the cardiotoxicity relating to the hERG-gene. The hERG residues interacting after 100 ns are LEU:697, THR:708, PHE:656, HIS:674, HIS:703, TRP:705 and ASN:709 and the hERG-ligand-16 complex trajectory showed stable behaviour with lesser fluctuations in the entire simulation of 200 ns.Communicated by Ramaswamy H. Sarma.

Nature and Strength of Sulfur-Centered Hydrogen Bond in Methanethiol Aqueous Solutions.

Methanethiol (M) and water (W) clusters like dimers (M1W1, M2, and W2), trimers (M1W2, M2W1, M3, and W3), and tetramers (M1W3, M2W2, M3W1, M4, and W4) were studied to assess the strength of sulfur-centered hydrogen bonding using different levels of theories, viz, HF, MP2, MP3, MP4, B3LYP, B3LYP-D3, CCSD, CCSD(T)-F12, and CCSD(T) along with aug-cc-pVNZ (where N = D, T, and Q) basis sets. Interaction energies were found to be in the range of -3.3 to -5.3 kcal/mol for the dimers, -8.0 to -16.7 kcal/mol for the trimers, and -13.5 to -29.5 kcal/mol for the tetramers at the B3LYP-D3/CBS limit level of theory. Normal modes of vibrations computed at the B3LYP/cc-pVDZ level of theory were seen to be in good agreement with the experimental values. Local energy decomposition calculations using the DLPNO-CCSD(T) level of theory indicated the domination of electrostatic interactions' contribution to the interaction energy in all cluster systems. Furthermore, atoms in molecules and natural bond orbital calculations both carried out at the B3LYP-D3/aug-cc-pVQZ level of theory aided in visualizing the hydrogen bonds besides proving a rationale for the strength of the hydrogen bonds and thereby the stability of these cluster systems.