Development of a reporter gene-based assay for the bioactivity determination of rhLH pharmaceutical products.

To establish an in vitro biological activity detection method for luteinizing hormone (LH), the hLHCGR-CREB-HEK293 cell line was constructed to stably express human luteinizing hormone/chorionic gonadotropin receptor (hLHCGR). After optimization, the rhLH starting working concentration was 800 mIU/mL with 4-fold serial dilutions, 10 concentrations and an incubation time of 5 h. The method was confirmed to be highly specific, with good accuracy, precision and linearity, meeting the needs of process research and release testing, and can be used as a routine detection method for LH biological activity. With the increasing demand for research and development of rhLH biologically similar drugs, establishing a stable and simple activity assay method to evaluate the biological activity of rhLH can provide technical support for quality control of rhLH products and powerful tools for comparability research of similar products.

Epilepsy as the symptom of a spinocerebellar ataxia 13 in a patient presenting with a mutation in the KCNC3 gene.

BACKGROUND: The spinocerebellar ataxias (SCAs) refer to a diverse group of neurodegenerative illnesses that vary clinically and genetically. One of the rare subtypes within this group is SCA13, caused by mutations in the KCNC3 gene. Currently, the prevalence of SCA13 remains uncertain, with only a couple of cases being documented in the Chinese population. This study presented a case study of SCA13, where the patient exhibited clinical symptoms of epilepsy and ataxia. The confirmation of the diagnosis was done through Whole Exome Sequncing. CASE PRESENTATION: Since childhood, the seventeen-year-old patient has not been capable of participating in numerous sporting activities and has experienced multiple episodes of unconsciousness within the last two years. The neurological evaluation showed a lack of coordination in the lower limbs. Cerebellar atrophy was detected through brain magnetic resonance imaging (MRI). The patient's gene detection results showed that they exhibit a heterozygous c.1268G > A mutation in the KCNC3 gene located at chr19:50826942. Antiepileptic treatment was promptly administered to the patient, and as a result, her epileptic seizures were resolved quickly. She has since remained free of seizures. After a one-year follow-up, there was no apparent improvement in the patient's health status except seizure free, which may have worsened. CONCLUSION: The case study highlights the importance of actively combining cranial MRI with genetic detection in patients with ataxia of no known cause, particularly in children and young patients, to establish an possibly obvious detection. Patients who are young and have ataxia that is first accompanied by extrapyramidal and epilepsy syndromes should be aware of the potential of having SCA13.

The efficacy and safety of intravenous tirofiban in the treatment of acute ischemic stroke patients with early neurological deterioration.

WHAT IS KNOWN AND OBJECTIVE: Many patients with acute ischemic stroke (AIS) develop early neurological deterioration (END), leading to disabilities or death. Thus, this study aimed to investigate the efficacy and safety of intravenous tirofiban in treating patients with AIS and END who missed the thrombolysis time window. METHODS: A total of 123 AIS-END patients participated in the study between January 2021 and December 2021. Patients were randomized into the tirofiban group (n = 63) and the control group (n = 60) based on whether a tirofiban injection was administered. The National Institute of Health Stroke Scale (NIHSS) was used to assess neurological function at the 48th hour and on the 7th day after intervention, and the modified Rankin Scale (mRS) was used to assess neurological recovery 90 days after AIS. Adverse reactions during the intervention were recorded for safety analysis. RESULTS AND DISCUSSION: The 7th day NIHSS and 90th day post-AIS mRS scores of the tirofiban group were significantly lower than those of the control group (p < 0.05), while the 90th day good prognosis (mRS ≤ 2) rate of the tirofiban group was significantly higher (84.13% vs. 65.00%, p < 0.05). Logistic regression demonstrated a protective effect of tirofiban for good prognosis in AIS patients with END (OR = 4.675, 95% CI [1.012-21.605], p < 0.05). No cases of intracranial haemorrhage transformation or death were observed during the treatment in either group. WHAT IS NEW AND CONCLUSION: Tirofiban injection exhibited a high safety profile and significantly improved the prognosis of AIS-END patients who missed the intravenous thrombolysis time window.

Pharmacological Treatment of Alzheimer's Disease: Insights from Drosophila melanogaster.

Aging is an ineluctable law of life. During the process of aging, the occurrence of neurodegenerative disorders is prevalent in the elderly population and the predominant type of dementia is Alzheimer's disease (AD). The clinical symptoms of AD include progressive memory loss and impairment of cognitive functions that interfere with daily life activities. The predominant neuropathological features in AD are extracellular ß-amyloid (Aß) plaque deposition and intracellular neurofibrillary tangles (NFTs) of hyperphosphorylated Tau. Because of its complex pathobiology, some tangible treatment can only ameliorate the symptoms, but not prevent the disease altogether. Numerous drugs during pre-clinical or clinical studies have shown no positive effect on the disease outcome. Therefore, understanding the basic pathophysiological mechanism of AD is imperative for the rational design of drugs that can be used to prevent this disease. Drosophila melanogaster has emerged as a highly efficient model system to explore the pathogenesis and treatment of AD. In this review we have summarized recent advancements in the pharmacological research on AD using Drosophila as a model species, discussed feasible treatment strategies and provided further reference for the mechanistic study and treatment of age-related AD.

Cerebral amyloid angiopathy-related inflammation: a case report presenting with a rare variant in SORL1 gene.

BACKGROUND: Cerebral amyloid angiopathy-related inflammation (CAA-ri) is a rare clinical entity, characterized by headaches, seizures, rapidly progressive cognitive decline, behavioral changes and magnetic resonance imaging (MRI) findings underlying the autoimmune and inflammatory reaction at the level of CAA-affected vessel. CAA-ri is likely responsive to corticosteroid. MRI shows asymmetric and multifocal white matter hyperintensity (WMH) lesions and multiple cerebral microbleeds. Apolipoprotein E (ApoE) ε4 homozygosity is associated with CAA-ri strongly [Neurology 68(17):1411-1416, 2007, Ann Neurol 73(4):449-458, 2013, J Alzheimers Dis 44(4):1069-1074, 2015]. SORL1 processes a causal involvement in Alzheimer's disease (AD) as a proposed modulator of the amyloid precursor protein (APP). It is unclear whether SORL1 is involved with CAA-ri or not. CASE PRESENTATION: A 48-year-old woman suffered from a one-day history of a headache, nausea, and vomiting. Neurological examination revealed normal. We diagnosed this case as probable CAA-ri according to the clinic manifestations and MRI. Gene detection indicated a rare variant in SORL1 and ApoE ε4 homozygosity. When treated with corticosteroid, the patient's clinical symptoms and MRI manifestations were almost relieved. However, when keeping the corticosteroid withdrawal for three months, the patient relapsed with a headache and typical images on MRI emerged. Corticosteroid therapy was effective again. Unfortunately, susceptibility weighted imaging (SWI) showed increased microbleeds. With tapering corticosteroid slowly, no recurrence was found on this patient with four-month follow-up. CONCLUSION: A variant of SORL1 may be associated with CAA-ri, recurrence of disease could be detected with MRI by an increased microbleeds. Our case report suggests that corticosteroid therapy might be effective for CAA-ri.

VhaAC39-1 regulates gut homeostasis and affects the health span in Drosophila.

Gut homeostasis is a dynamically balanced state to maintain intestinal health. Vacuolar ATPases (V-ATPases) are multi-subunit proton pumps that were driven by ATP hydrolysis. Several subunits of V-ATPases may be involved in the maintenance of intestinal pH and gut homeostasis in Drosophila. However, the specific role of each subunit in this process remains to be elucidated. Here, we knocked down the Drosophila gene VhaAC39-1 encoding the V0d1 subunit of V-ATPases to assess its function in gut homeostasis. Knockdown of VhaAC39-1 resulted in the loss of midgut acidity, the increase of the number of gut microbiota and the impairment of intestinal epithelial integrity in flies. The knockdown of VhaAC39-1 led to the hyperproliferation of intestinal stem cells, increasing the number of enteroendocrine cells, and activated IMD signaling pathway and JAK-STAT signaling pathway, inducing intestinal immune response of Drosophila. In addition, knockdown of VhaAC39-1 caused the disturbance of many physiological indicators such as food intake, triglyceride level and fecundity of flies, which ultimately led to the shortening of the life span of Drosophila. These results shed light on the gut homeostasis mechanisms which would help to identify interventions to promote healthy aging.

Effects of anti-aging interventions on intestinal microbiota.

Identifying ways to deal with the challenges presented by aging is an urgent task, as we are facing an aging society. External factors such as diet, exercise and drug therapy have proven to be major elements in controlling healthy aging and prolonging life expectancy. More recently, the intestinal microbiota has also become a key factor in the anti-aging process. As the intestinal microbiota changes with aging, an imbalance in intestinal microorganisms can lead to many age-related degenerative diseases and unhealthy aging. This paper reviews recent research progress on the relationship between intestinal microorganisms and anti-aging effects, focusing on the changes and beneficial effects of intestinal microorganisms under dietary intervention, exercise and drug intervention. In addition, bacteriotherapy has been used to prevent frailty and unhealthy aging. Most of these anti-aging approaches improve the aging process and age-related diseases by regulating the homeostasis of intestinal flora and promoting a healthy intestinal environment. Intervention practices based on intestinal microorganisms show great potential in the field of anti-aging medicine.

Comparative proteomics analysis of dietary restriction in Drosophila.

To explore the underlying mechanism of dietary restriction (DR) induced lifespan extension in fruit flies at protein level, we performed proteome sequencing in Drosophila at day 7 (young) and day 42 (old) under DR and ad libitum (AL) conditions. A total of 18629 unique peptides were identified in Uniprot, corresponding to 3,662 proteins. Among them, 383 and 409 differentially expressed proteins (DEPs) were identified from comparison between DR vs AL at day 7 and 42, respectively. Bioinformatics analysis revealed that membrane-related processes, post-transcriptional processes, spliceosome and reproduction related processes, were highlighted significantly. In addition, expression of proteins involved in pathways such as spliceosomes, oxidative phosphorylation, lysosomes, ubiquitination, and riboflavin metabolism was relatively higher during DR. A relatively large number of DEPs were found to participate in longevity and age-related disease pathways. We identified 20 proteins that were consistently regulated during DR and some of which are known to be involved in ageing, such as mTORC1, antioxidant, DNA damage repair and autophagy. In the integration analysis, we found 15 genes that were stably regulated by DR at both transcriptional as well as translational levels. Our results provided a useful dataset for further investigations on the mechanism of DR and aging.

lncRNA ZEB1-AS1 Mediates Oxidative Low-Density Lipoprotein-Mediated Endothelial Cells Injury by Post-transcriptional Stabilization of NOD2.

Oxidized-low density lipoprotein (ox-LDL) can induce injury of endothelial cells, causing atherosclerosis, which is an important initial event in several cardiovascular diseases. Long non-coding RNAs (lncRNAs) have emerged as regulators of diverse biological processes, but their specific biological functions and biochemical mechanisms in ox-LDL-induced endothelial cell injury have not been well investigated. Here, we describe the initial functional analysis of a poorly characterized human lncRNA ZEB1 antisense 1 (ZEB1-AS1). We found that ox-LDL treatment could induce a decreased cell viability and an increased cell apoptosis in endothelial cells, and knockdown of ZEB1-AS1 significantly reversed this effect. Mechanistically, ox-LDL treatment could sequester p53 from binding to ZEB1-AS1 promoter region, causing transcriptional activation and upregulation of ZEB1-AS1. Moreover, enhanced ZEB1-AS1 could upregulate Nucleotide-Binding Oligomerization Domain 2 (NOD2) expression through recruiting leucine-rich pentatricopeptide repeat motif-containing protein (LRPPRC) to stabilize NOD2 mRNA. Experimental data showed that knockdown of NOD2 or LRPPRC dramatically abrogated the functional role of ZEB1-AS1 in ox-LDL-induced endothelial cell injury. In summary, we demonstrated that lncRNA ZEB1-AS1 regulates the ox-LDL-induced endothelial cell injury via an LRPPRC-dependent mRNA stabilization mechanism. Therefore, ZEB1-AS1 may serve as a multi-potency target to overcome endothelial cell injury, atherosclerosis and other cardiovascular diseases.

Transorbital Doppler with carotid siphon monitoring detects right-to-left shunt effectively.

Background Transtemporal Doppler (TTD) with middle cerebral artery (MCA) is widely used for right-to-left shunt (RLS) detection. However, an alternative method for patients without suitable temporal bone windows should be established. The present study prospectively evaluated the effectiveness of transorbital Doppler (TOD) with carotid siphon (CS) monitoring in detecting RLS. Methods A total of 357 subjects with sufficient temporal bone windows underwent simultaneous TTD with MCA and TOD with CS. After injection of microbubbles, the numbers of artificial high-intensity signals were recorded at rest and after Valsalva maneuver. Results TOD with CS detected RLS in 146 patients. Sensitivity was 97.1%, specificity 95%, positive predictive value 92.5%, and negative predictive value 98.1%. The total positive rates for RLS detection by CS (40.9%) and MCA (37.8%) monitoring were comparable without significant difference, but TOD with CS detected significantly more grade 2 and 3 RLS than TTD with MCA (p = 0.001). The RLS rates of cryptogenic stroke patients was significantly higher than that of healthy controls, and RLS in cryptogenic stroke was remarkably higher than that in transient ischemia attack patients (p < 0.05). TOD with CS examined significantly more grade 2 and 3 RLSs than the MCA approach in the cryptogenic stroke patients (p = 0.037). Conclusion TOD with CS monitoring is able to detect RLS effectively in different populations including healthy subjects, cryptogenic stroke, transient ischemia attack, and migraine patients. In comparing to the TTD with MCA approach, TOD with CS monitoring could detect comparable rate of RLS, but more high grades of RLS.