Antiretroviral therapy adherence among peripartum women with HIV in Kenya: an explanatory mixed methods study using dry blood spot measures and narrative interviews.

ABSTRACTAdherence to antiretroviral therapy (ART) remains sub-optimal among pregnant and postpartum women with HIV (PPWH) in high HIV prevalence low resource settings with few effective behavioral interventions. A large body of qualitative literature has established general barriers and facilitators to ART adherence in PPWH at various levels (individual, interpersonal, structural). However, research exploring the underlying behavioral mechanisms of ART adherence in PPWH with objectively verified adherence biomarkers is extremely limited. We conducted 24 in-depth interviews with postpartum women in western Kenya who had linked ART drug concentrations obtained from three dried blood spot samples across the peripartum period. Among PPWH with a low drug concentration (n = 13) compared to those with continuously high drug concentrations (n = 11), distinct themes emerged related to HIV status disclosure, social support, interactions with the health system, and health beliefs. By combining ART biomarkers with patient reported challenges, there is the potential for real-time interventions to support sustained ART adherence among PPWH and improve maternal and infant health outcomes.

DNA damage targeted therapy for advanced breast cancer.

Breast cancer (BC) is the most prevalent malignancy affecting women worldwide, including Portugal. While the majority of BC cases are sporadic, hereditary forms account for 5-10% of cases. The most common inherited mutations associated with BC are germline mutations in the BReast CAncer (BRCA) 1/2 gene (gBRCA1/2). They are found in approximately 5-6% of BC patients and are inherited in an autosomal dominant manner, primarily affecting younger women. Pathogenic variants within BRCA1/2 genes elevate the risk of both breast and ovarian cancers and give rise to distinct clinical phenotypes. BRCA proteins play a key role in maintaining genome integrity by facilitating the repair of double-strand breaks through the homologous recombination (HR) pathway. Therefore, any mutation that impairs the function of BRCA proteins can result in the accumulation of DNA damage, genomic instability, and potentially contribute to cancer development and progression. Testing for gBRCA1/2 status is relevant for treatment planning, as it can provide insights into the likely response to therapy involving platinum-based chemotherapy and poly[adenosine diphosphate (ADP)-ribose] polymerase inhibitors (PARPi). The aim of this review was to investigate the impact of HR deficiency in BC, focusing on BRCA mutations and their impact on the modulation of responses to platinum and PARPi therapy, and to share the experience of Unidade Local de Saúde Santa Maria in the management of metastatic BC patients with DNA damage targeted therapy, including those with the Portuguese c.156_157insAlu BRCA2 founder mutation.

Patterns of daily oral HIV PrEP adherence among people who inject drugs in Ukraine: an analysis of biomarkers.

INTRODUCTION: Daily oral HIV pre-exposure prophylaxis (PrEP) with tenofovir/emtricitabine (TDF/FTC) is recommended for people who inject drugs (PWID) but coverage is low. The real-life effectiveness of PrEP among PWID is unknown as previous studies were conducted in controlled settings and mainly relied on self-report. Analysis of PrEP metabolites-tenofovir diphosphate (TFVdp) and emtricitabine triphosphate (FTCtp)-offers an objective measure of adherence. METHODS: To analyse longitudinal patterns of PrEP adherence among PWID in Ukraine, we used data from a community-based implementation trial conducted in Kyiv between July 2020 and March 2021 to test the efficacy of SMS reminders to improve adherence. Among 199 enrolled participants, 156 (78.4%) were retained through 6 months. Based on TFVdp/FTCtp levels assessed at 3 and 6 months, we identified groups with various adherence patterns (adherent at ≥2 doses/week, improved, worsened, non-adherent). Correlates of adherence were analysed using multinomial logistic regression. RESULTS: Most participants (53.8%, n = 84/156) had no detectable metabolites at both assessments; 7.1% (n = 11/156) were consistently taking ≥2 doses/week; 1.3% (n = 2/156) were consistently taking ≥4 doses/week; 13.5% (n = 21/156) exhibited improved and 21.8% (n = 34/156) had worsened adherence at 6 compared to 3 months. "White coat compliance" (increased dosing prior to assessment) was common. Consistent adherence was associated with SMS reminders, younger age, employment, lower income, longer injection drug use duration, recent high-risk injecting (receptive syringe sharing, using pre-filled syringe, back- or front-loading, container sharing), absence of overdose in the past 6 months, perceived HIV risk through sexual intercourse and higher PrEP self-efficacy. Alcohol consumption was associated with inconsistent PrEP use. Groups with improved and worsened adherence did not differ. CONCLUSIONS: Daily oral PrEP may not achieve the desired effectiveness among PWID as a standalone intervention, calling for testing of alternative PrEP formulations and innovative integrated risk reduction strategies, especially in the context of HIV epidemics associated with injection drug use in eastern Europe and central Asia and the public health crisis in Ukraine caused by the war with Russia. SMS reminders may be effective among PWID who prioritize PrEP. Our findings offer practical guidance in identifying PWID who are likely to benefit from PrEP and those who need additional support.

P2Y12 adenosine receptor inhibitors preloading among patients with non-ST-elevation acute coronary syndromes; insights from a nationwide multicenter registry.

BACKGROUND: The safety and efficacy of treatment with P2Y12 adenosine-diphosphate receptor inhibitors (P2Y12-RI) before coronary angiography among patients with non-ST-segment elevation acute coronary syndromes (NSTEACS) are questionable. AIMS: To assess the pretreatment rate with P2Y12-RI and its association with ischemic and bleeding risks among patients with NSTEACS. METHODS: The study comprised patients with NSTEACS referred for coronary angiography and included in the Acute Coronary Syndrome Israeli Surveys between 2013 and 2021. Patients were divided into two groups according to the timing of P2Y12-RI loading concerning coronary angiography: pretreatment and posttreatment. The primary endpoints were 30-day major adverse cardiovascular events (MACE; composite of cardiovascular mortality, myocardial infarction, stroke, stent thrombosis, and urgent revascularization) and 1-year all-cause mortality. RESULTS: Of 3076 patients, 2423 (78.8%) received pretreatment with a P2Y12-RI, and 653 (21.2%) received P2Y12-RI posttreatment. Prasugrel and ticagrelor were used more in the posttreatment group compared to the pretreatment group (16% vs. 6% and 38% vs. 25%, respectively, p < 0.001 for both). No difference was observed in the rate of 30-day MACE comparing pretreatment and posttreatment (5.3% vs. 2.2%, respectively, p = 0.62). A sensitivity analysis of 30-day MACE among patients from the 2021 survey demonstrated similar results (2.5% in the posttreatment group vs. 8.0% in the pretreatment group, p = 0.13). There were no differences in 1-year all-cause mortality rates between the pretreatment and posttreatment groups (4.8% vs. 3.8%, p = 0.31). CONCLUSIONS: Among patients with NSTEACS referred for an invasive strategy, the P2Y12-RI posttreatment strategy was associated with similar 30-day and 1-year MACE as the pretreatment strategy. These large-scale, multicenter, real-world data provide reassurance on the safety and efficacy of delaying P2Y12-IR until after coronary stratification to improve clinical decision-making.

Screening of ent-copalyl diphosphate synthase and metabolic engineering to achieve de novo biosynthesis of ent-copalol in Saccharomyces cerevisiae.

The diterpene ent-copalol is an important precursor to the synthesis of andrographolide and is found only in green chiretta (Andrographis paniculata). De novo biosynthesis of ent-copalol has not been reported, because the catalytic activity of ent-copalyl diphosphate synthase (CPS) is very low in microorganisms. In order to achieve the biosynthesis of ent-copalol, Saccharomyces cerevisiae was selected as the chassis strain, because its endogenous mevalonate pathway and dephosphorylases could provide natural promotion for the synthesis of ent-copalol. The strain capable of synthesizing diterpene geranylgeranyl pyrophosphate was constructed by strengthening the mevalonate pathway genes and weakening the competing pathway. Five full-length ApCPSs were screened by transcriptome sequencing of A. paniculata and ApCPS2 had the best activity and produced ent-CPP exclusively. The peak area of ent-copalol was increased after the ApCPS2 saturation mutation and its configuration was determined by NMR and ESI-MS detection. By appropriately optimizing acetyl-CoA supply and fusion-expressing key enzymes, 35.6 mg/L ent-copalol was generated. In this study, de novo biosynthesis and identification of ent-copalol were achieved and the highest titer ever reported. It provides a platform strain for the further pathway analysis of andrographolide and derivatives and provides a reference for the synthesis of other pharmaceutical intermediates.

Assessment of Erythrocyte Transketolase, Whole Blood Thiamine Diphosphate, and Human Milk Thiamine Concentrations to Identify Infants and Young Children Responding Favorably to Therapeutic Thiamine Administration: Findings from the Lao Thiamine Study, a Prospective Cohort Study.

Background: There is limited information on relationships among biomarkers of thiamine status (whole blood thiamine diphosphate [ThDP], erythrocyte transketolase activity coefficient [ETKac], and human milk thiamine [MTh]) and clinical manifestations of thiamine deficiency. Objectives: This study aimed to explore correlations among these biomarkers and thiamine responsive disorders (TRDs), a diagnosis based on favorable clinical response to thiamine. Methods: Hospitalized infants and young children (aged 21 d to <18 mo) with respiratory, cardiac, and/or neurological symptoms suggestive of thiamine deficiency were treated with parenteral thiamine (100 mg daily) for ≥3 d alongside other treatments and re-examined systematically. Clinical case reports were reviewed by 3 pediatricians, who determined TRD or non-TRD status. Children in a community comparison group were matched by age, sex, and residence. Venous whole blood ThDP and MTh were determined by high-performance liquid chromatography fluorescence detection and ETKac in washed erythrocytes by ultraviolet spectrophotometry. Associations between biomarkers were assessed using Spearman correlations, and biomarker cutoffs predictive of TRD and ETKac >1.25 were explored using area under the receiver operating characteristic curve framework. Results: Thiamine biomarkers were available for 287 hospitalized children and 228 community children (mean age 4.7 mo; 59.4% male). Median (interquartile range [IQR]) ThDP and ETKac were 66.9 nmol/L (IQR: 41.4, 96.9 nmol/L) and 1.25 nmol/L (IQR: 1.11, 1.48 nmol/L), respectively, among hospitalized children, and 64.1 nmol/L (IQR: 50.0, 85.3 nmol/L) and 1.22 nmol/L (IQR: 1.12, 1.37 nmol/L) among 228 community children (P > 0.05 for both). Forty-five percent of breastfeeding mothers of infants <6 mo had MTh <90 µg/L. ThDP and ETKac, but not MTh, were significantly different between 152 children with TRD and 122 without TRD, but overlapping distributions undermined prediction of individual responses to thiamine. Conclusions: Although ETKac, ThDP, and MTh are useful biomarkers of population thiamine status, none of the biomarkers reliably identified individual children with TRD. ThDP is more practical for population assessment because preparing washed erythrocytes is not required.This trial was registered at clinicaltrials.gov as NCT03626337.

Transcriptome analysis revealed the role of moderate exogenous methyl jasmonate treatments in enhancing the metabolic pathway of L-borneol in the Blumea balsamifera.

Introduction: Blumea balsamifera L. (Ainaxiang) DC. is a perennial herb of the compositae family. It is also the primary source of natural borneol. Endo-borneol, the principal medical active element in B. balsamifera, is anti-inflammatory, antioxidant, and analgesic; enhances medicine absorption; refreshes; and is used as a spice and in cosmetic. Industrialization of B. balsamifera is limited by its low L-borneol concentration. Thus, understanding the accumulation pattern of the secondary metabolite endo-borneol and its synthesis process in secondary metabolism is critical for increasing B. balsamifera active ingredient content and cultivation efficiency. Methods: In this work, B. balsamifera was treated with varying concentrations (1.00 and 10.00 mmol/L) of methyl jasmonate (MeJA) as an exogenous foliar activator. The physiological parameters and L-borneol concentration were then assessed. Transcriptome sequencing of B. balsamifera-induced leaves was used to identify key genes for monoterpene synthesis. Results: The treatment effect of 1 mmol/L MeJA was the best, and the leaves of all three leaf positions accumulated the highest L-borneol after 120 h, correspondingly 3.043 mg·g-1 FW, 3.346 mg·g-1 FW, and 2.044 mg·g-1 FW, with significant differences from the control. The main assembly produced 509,285 transcripts with min and max lengths of 201 and 23,172, respectively. DEG analysis employing volcano blots revealed 593, 224, 612, 2,405, 1,353, and 921 upregulated genes and 4, 123, 573, 1,745, 766, and 763 downregulated genes in the treatments D1_1vsCK, D1_10vsCK, D2_1vsCK, D2_10vsCK, D5_1vsCK, and D5_10vsCK. Interestingly, when exposed to MeJA treatments, the MEP pathway's unigenes express themselves more than those of the MVA route. Finally, when treated with 1 mmol/L, the genes DXR, DXS, and GPS showed increased expression over time. At the same time, a 10 mmol/L therapy resulted in elevated levels of ispH and GGPS. Discussion: Our preliminary research indicates that exogenous phytohormones can raise the level of L borneol in B. balsamifera (L.) DC when given in the appropriate amounts. The most significant discovery made while analyzing the effects of different hormones and concentrations on B. balsamifera (L.) DC was the effect of 1 mmol/L MeJA treatment.

Synthesis and evaluation of (E)-4-hydroxy-3-methyl-but-2-enyl diphosphate analogs as competitive partial agonists of butyrophilin 3A1.

Phosphoantigens (pAgs) induce conformational changes after binding to the intracellular region of BTN3A1 which result in its clustering with BTN2A1, forming an activating ligand for the Vγ9Vδ2 T cell receptor. Here, we designed a small panel of bulky analogs of the prototypical pAg (E)-4-hydroxy-3-methyl-but-2-enyl diphosphate (HMBPP) that contain an aromatic ring attached to the C-3 position in place of methyl group. These compounds bind with high affinity to BTN3A1 but fail to fully support its interaction with BTN2A1 and only partially trigger T cell activation relative to HMBPP. Furthermore, they can compete with HMBPP for cellular binding to BTN3A1 and reduce the cellular response to HMBPP, a classic partial agonist phenotype. Trifluoromethyl analog 6e was the weakest agonist but the strongest inhibitor of HMBPP ELISA response. Our study provides a rationale for the mode of action of pAg-induced γδ T cell activation and provides insights into other naturally occurring BTN proteins and their respective ligands.

Abnormal metabolism in melanocytes participates in the activation of dendritic cell in halo nevus.

A comprehensive analysis of spatial transcriptomics was carried out to better understand the progress of halo nevus. We found that halo nevus was characterized by overactive immune responses, triggered by chemokines and dendritic cells (DCs), T cells, and macrophages. Consequently, we observed abnormal cell death, such as apoptosis and disulfidptosis in halo nevus, some were closely related to immunity. Interestingly, we identified aberrant metabolites such as uridine diphosphate glucose (UDP-G) within the halo nevus. UDP-G, accompanied by the infiltration of DCs and T cells, exhibited correlations with certain forms of cell death. Subsequent experiments confirmed that UDP-G was increased in vitiligo serum and could activate DCs. We also confirmed that oxidative response is an inducer of UDP-G. In summary, the immune response in halo nevus, including DC activation, was accompanied by abnormal cell death and metabolites. Especially, melanocyte-derived UDP-G may play a crucial role in DC activation.

Safety Assessment of Polyol Phosphates as Used in Cosmetics.

The Expert Panel for Cosmetic Ingredient Safety (Panel) reviewed the safety of 10 polyol phosphates. Some of the possible functions in cosmetics that are reported for this ingredient group are chelating agents, oral care agents, and skin conditioning agents. The Panel reviewed relevant data relating to the safety of these ingredients under the intended conditions of use in cosmetic formulations, and concluded that Sodium Phytate, Phytic Acid, Phytin, and Trisodium Inositol Triphosphate are safe in cosmetics in the present practices of use and concentration described in the safety assessment. The Panel also concluded that the data are insufficient to determine the safety of the following 6 ingredients as used in cosmetics: Disodium Glucose Phosphate, Manganese Fructose Diphosphate, Sodium Mannose Phosphate, Trisodium Fructose Diphosphate, Xylityl Phosphate, and Zinc Fructose Diphosphate.

Intraocular kinetics of pathological ATP after photoreceptor damage in rhegmatogenous retinal detachment.

PURPOSE: Extracellular Adenosine triphosphate (ATP) released by dying cells may cause a secondary cell death in neighboring cells in retinal degeneration. We investigated intraocular ATP kinetics to gain mechanical insights into the pathology in rhegmatogenous retinal detachment (RRD). STUDY DESIGN: Retrospective clinical study. METHODS: Vitreous or subretinal fluids (SRF) were obtained from patients with RRD (n=75), macular hole (MH; n=20), and epiretinal membrane (ERM; n=35) during vitrectomy. ATP levels in those samples were measured by luciferase assay. RESULTS: Mean ATP levels in the vitreous from RRD patients were significantly higher compared to those from MH and ERM patients (2.3 and 0.3 nM, respectively. P<0.01). Mean ATP levels in the SRF from RRD (11.7 nM) were higher than those in the vitreous from RRD (P<0.01). Mean ATP levels in the vitreous with short durations (1-8 days) of RRD were higher compared to those with long durations (>8 days) (3.2 and 1.4 nM, respectively. P<0.05). Similarly, ATP in SRF with short durations were higher than those with long durations (23.8 and 3.6 nM, respectively. P<0.05). Furthermore, the concentrations of ectonucleoside triphosphate diphosphohydrolase-1 (ENTPD1), a major ATP degradative enzyme, in the vitreous from RRD were higher than those from MH/ERM (1.2 and 0.2 ng/ml, respectively. P<0.01). ENTPD1 expression was localized in the cytoplasm of CD11b-positive infiltrating cells in the vitreous and retinal cells. CONCLUSION: ATP increased in the vitreous and SRF in RRD and decreased over time with an upregulation of ENTPD1. The kinetics indicate the pathological mechanism of the excessive extracellular ATP after RRD.

Increased Expression of Mevalonate Pathway-Related Enzymes in Angiotensin II-Induced Abdominal Aortic Aneurysms.

Abdominal aortic aneurysm (AAA) is characterized by permanent luminal expansion and a high mortality rate due to aortic rupture. Despite the identification of abnormalities in the mevalonate pathway (MVA) in many diseases, including cardiovascular diseases, the potential impact of this pathway on AAA remains unclear. This study aims to investigate whether the expression of the MVA-related enzyme is altered during the progression of angiotensin II (Ang II) -induced AAA.Ang II 28D and Ang II 5D groups were continuously perfused with Ang II for 28 days and 5 days, respectively, and the Sham group was perfused with saline. The general and remodeling characteristics of AAA were determined by biochemical and histological analysis. Alteration of MVA-related enzyme expressions was revealed by western blot and single-cell RNA sequencing (scRNA-seq).The continuous Ang II infusion for 28 days showed significant aorta expansion and arterial remodeling. Although the arterial diameter slightly increased, the aneurysm formation was not found in Ang II induction for 5 days. MVA-related enzyme expression and activation of small GTP-binding proteins were significantly increased after Ang II-induced. As verified by scRNA-seq, the key enzyme gene expression was also higher in Ang II 28D. Similarly, it was detected that the expression levels of the above enzymes and the activity of small G proteins were elevated in the early stage of AAA as induced by Ang II infusion for 5 days.Continuous Ang II infusion-induced abdominal aortic expansion and arterial remodeling were accompanied by altered expression of key enzymes in the MVA.

High cost of chemotherapy for gynecologic malignancies.

BACKGROUND: The prognosis of gynecological malignancies has improved with the recent advent of molecularly targeted drugs and immune checkpoint inhibitors. However, these drugs are expensive and contribute to the increasing costs of medical care. METHODS: The Japanese Clinical Oncology Group (JCOG) Health Economics Committee conducted a questionnaire survey of JCOG-affiliated facilities from July 2021 to June 2022 to assess the prevalence of high-cost regimens. RESULTS: A total of 57 affiliated facilities were surveyed regarding standard regimens for advanced ovarian and cervical cancers for gynecological malignancies. Responses were obtained from 39 facilities (68.4%) regarding ovarian cancer and 37 (64.9%) concerning cervical cancer, with respective case counts of 854 and 163. For ovarian cancer, 505 of 854 patients (59.1%) were treated with regimens that included PARP inhibitors, costing >500 000 Japanese yen monthly, while 111 patients (13.0%) received treatments that included bevacizumab, with costs exceeding 200 000 Japanese yen monthly. These costs are ~20 and ~10 times higher than those of the conventional regimens, respectively. For cervical cancer, 79 patients (48.4%) were treated with bevacizumab regimens costing >200 000 Japanese yen per month, ~10 times the cost of conventional treatments. CONCLUSIONS: In this survey, >70% of patients with ovarian cancer were treated with regimens that included poly (adenosine diphosphate-ribose) polymerase (PARP) inhibitors or bevacizumab; ~50% of patients with cervical cancer were treated with regimens containing bevacizumab. These treatments were ~10 and ~20 times more expensive than conventional regimens, respectively. These findings can inform future health economics studies, particularly in assessing cost-effectiveness and related matters.

Nucleoside Diphosphate Kinases Are ATP-Regulated Carriers of Short-Chain Acyl-CoAs.

Nucleoside diphosphate (NDP) kinases 1 and 2 (NME1/2) are well-characterized enzymes known for their NDP kinase activity. Recently, these enzymes have been shown by independent studies to bind coenzyme A (CoA) or acyl-CoA. These findings suggest a hitherto unknown role for NME1/2 in the regulation of CoA/acyl-CoA-dependent metabolic pathways, in tight correlation with the cellular NTP/NDP ratio. Accordingly, the regulation of NME1/2 functions by CoA/acyl-CoA binding has been described, and additionally, NME1/2 have been shown to control the cellular pathways consuming acetyl-CoA, such as histone acetylation and fatty acid synthesis. NME1/2-controlled histone acetylation in turn mediates an important transcriptional response to metabolic changes, such as those induced following a high-fat diet (HFD). This review discusses the CoA/acyl-CoA-dependent NME1/2 activities and proposes that these enzymes be considered as the first identified carriers of CoA/short-chain acyl-CoAs.

CDP-DAG synthesis by peripheral membrane-bound Tam41-type enzymes.

Cytidine diphosphate diacylglycerol (CDP-DAG) is a critical intermediate that is converted to multiple phospholipids in prokaryotes and eukaryotes. In budding yeast, CDP-DAG synthesis from cytidine triphosphate (CTP) and phosphatidic acid (PA) is catalyzed by the membrane-integrated protein Cds1 in the endoplasmic reticulum and the peripheral membrane-bound protein Tam41 in mitochondria. Although a recent study revealed that the fission yeast SpTam41 consists of a nucleotidyltransferase domain and a winged helix domain, forming an active-site pocket for CTP binding between the two domains together with a C-terminal amphipathic helix for membrane association, how CTP and Mg2+, a most-favored divalent cation, are accommodated with PA remains obscure. A more recent report by Kimura et al. (J. Biochem. 2022; 171:429-441) solved the crystal structure of FbTam41, a functional ortholog from a Firmicutes bacterium, with CTP-Mg2+, successfully providing a detailed molecular view of CDP-DAG synthesis. In this commentary, our current understanding of Tam41-mediated reaction is discussed.

Significance of Ribonucleoside-diphosphate Reductase Subunit M2 in Lung Adenocarcinoma.

INTRODUCTION: The Ribonucleoside-diphosphate Reductase subunit M2 (RRM2) is known to be overexpressed in various cancers, though its specific functional implications remain unclear. This aims to elucidate the role of RRM2 in the progression of Lung Adenocarcinoma (LUAD) by exploring its involvement and potential impact. METHODS: RRM2 data were sourced from multiple databases to assess its diagnostic and prognostic significance in LUAD. We evaluated the association between RRM2 expression and immune cell infiltration, analyzed its function, and explored the effects of modulating RRM2 expression on LUAD cell characteristics through laboratory experiments. RESULTS: RRM2 was significantly upregulated in LUAD tissues and cells compared to normal counterparts (p<0.05), with rare genetic alterations noted (approximately 2%). This overexpression clearly distinguished LUAD from normal tissue (area under the curve (AUC): 0.963, 95% confidence intervals (CI): 0.946-0.981). Elevated RRM2 expression was significantly associated with adverse clinicopathological characteristics and poor prognosis in LUAD patients. Furthermore, a positive association was observed between RRM2 expression and immune cell infiltration. Pathway analysis revealed a critical connection between RRM2 and the cell cycle signaling pathway within LUAD. Targeting RRM2 inhibition effectively suppressed LUAD cell proliferation, migration, and invasion while promoting apoptosis. This intervention also modified the expression of several crucial proteins, including the downregulation of CDC25A, CDC25C, RAD1, Bcl-2, and PPM1D and the upregulation of TP53 and Bax (p < 0.05). CONCLUSION: Our findings highlight the potential utility of RRM2 expression as a biomarker for diagnosing and predicting prognosis in LUAD, shedding new light on the role of RRM2 in this malignancy.

Studying the impact of titanium dioxide nanoparticles on the expression of pivotal genes related to menthol biosynthesis and certain biochemical parameters in peppermint plants (Mentha Piperita L.).

BACKGROUND: This study examines the impact of titanium dioxide nanoparticles (TiO2NPs) on gene expression associated with menthol biosynthesis and selected biochemical parameters in peppermint plants (Mentha piperita L.). Menthol, the active ingredient in peppermint, is synthesized through various pathways involving key genes like geranyl diphosphate synthase, menthone reductase, and menthofuran synthase. Seedlings were treated with different concentrations of TiO2NPs (50, 100, 200, and 300 ppm) via foliar spray. After three weeks of treatment, leaf samples were gathered and kept at -70 °C for analysis. RESULTS: According to our findings, there was a significant elevation (P ≤ 0.05) in proline content at concentrations of 200 and 300 ppm in comparison with the control. Specifically, the highest proline level was registered at 200 ppm, reaching 259.64 ± 33.33 µg/g FW. Additionally, hydrogen peroxide and malondialdehyde content exhibited a decreasing trend following nanoparticle treatments. Catalase activity was notably affected by varying TiO2NP concentrations, with a significant decrease observed at 200 and 300 ppm compared to the control (P ≤ 0.05). Conversely, at 100 ppm, catalase activity significantly increased (11.035 ± 1.12 units/mg of protein/min). Guaiacol peroxidase activity decreased across all nanoparticle concentrations. Furthermore, RT-qPCR analysis indicated increased expression of the studied genes at 300 ppm concentration. CONCLUSIONS: Hence, it can be inferred that at the transcript level, this nanoparticle exhibited efficacy in influencing the biosynthetic pathway of menthol.

Glycolytic PFKFB3 and Glycogenic UGP2 Axis Regulates Perfusion Recovery in Experimental Hind Limb Ischemia.

BACKGROUND: Despite being in an oxygen-rich environment, endothelial cells (ECs) use anaerobic glycolysis (Warburg effect) as the primary metabolic pathway for cellular energy needs. PFKFB (6-phosphofructo-2-kinase/fructose-2,6-biphosphatase)-3 regulates a critical enzymatic checkpoint in glycolysis and has been shown to induce angiogenesis. This study builds on our efforts to determine the metabolic regulation of ischemic angiogenesis and perfusion recovery in the ischemic muscle. METHODS: Hypoxia serum starvation (HSS) was used as an in vitro peripheral artery disease (PAD) model, and hind limb ischemia by femoral artery ligation and resection was used as a preclinical PAD model. RESULTS: Despite increasing PFKFB3-dependent glycolysis, HSS significantly decreased the angiogenic capacity of ischemic ECs. Interestingly, inhibiting PFKFB3 significantly induced the angiogenic capacity of HSS-ECs. Since ischemia induced a significant in PFKFB3 levels in hind limb ischemia muscle versus nonischemic, we wanted to determine whether glucose bioavailability (rather than PFKFB3 expression) in the ischemic muscle is a limiting factor behind impaired angiogenesis. However, treating the ischemic muscle with intramuscular delivery of D-glucose or L-glucose (osmolar control) showed no significant differences in the perfusion recovery, indicating that glucose bioavailability is not a limiting factor to induce ischemic angiogenesis in experimental PAD. Unexpectedly, we found that shRNA-mediated PFKFB3 inhibition in the ischemic muscle resulted in an increased perfusion recovery and higher vascular density compared with control shRNA (consistent with the increased angiogenic capacity of PFKFB3 silenced HSS-ECs). Based on these data, we hypothesized that inhibiting HSS-induced PFKFB3 expression/levels in ischemic ECs activates alternative metabolic pathways that revascularize the ischemic muscle in experimental PAD. A comprehensive glucose metabolic gene qPCR arrays in PFKFB3 silenced HSS-ECs, and PFKFB3-knock-down ischemic muscle versus respective controls identified UGP2 (uridine diphosphate-glucose pyrophosphorylase 2), a regulator of protein glycosylation and glycogen synthesis, is induced upon PFKFB3 inhibition in vitro and in vivo. Antibody-mediated inhibition of UGP2 in the ischemic muscle significantly impaired perfusion recovery versus IgG control. Mechanistically, supplementing uridine diphosphate-glucose, a metabolite of UGP2 activity, significantly induced HSS-EC angiogenic capacity in vitro and enhanced perfusion recovery in vivo by increasing protein glycosylation (but not glycogen synthesis). CONCLUSIONS: Our data present that inhibition of maladaptive PFKFB3-driven glycolysis in HSS-ECs is necessary to promote the UGP2-uridine diphosphate-glucose axis that enhances ischemic angiogenesis and perfusion recovery in experimental PAD.

Preliminary X-ray diffraction and ligand-binding analyses of the N-terminal domain of hypothetical protein Rv1421 from Mycobacterium tuberculosis H37Rv.

Mycobacterium tuberculosis can reside and persist in deep tissues; latent tuberculosis can evade immune detection and has a unique mechanism to convert it into active disease through reactivation. M. tuberculosis Rv1421 (MtRv1421) is a hypothetical protein that has been proposed to be involved in nucleotide binding-related metabolism in cell-growth and cell-division processes. However, due to a lack of structural information, the detailed function of MtRv1421 remains unclear. In this study, a truncated N-terminal domain (NTD) of MtRv1421, which contains a Walker A/B-like motif, was purified and crystallized using PEG 400 as a precipitant. The crystal of MtRv1421-NTD diffracted to a resolution of 1.7 Šand was considered to belong to either the C-centered monoclinic space group C2 or the I-centered orthorhombic space group I222, with unit-cell parameters a = 124.01, b = 58.55, c = 84.87 Å, ß = 133.12° or a = 58.53, b = 84.86, c = 90.52 Å, respectively. The asymmetric units of the C2 or I222 crystals contained two or one monomers, respectively. In terms of the binding ability of MtRv1421-NTD to various ligands, uridine diphosphate (UDP) and UDP-N-acetylglucosamine significantly increased the melting temperature of MtRv1421-NTD, which indicates structural stabilization through the binding of these ligands. Altogether, the results reveal that a UDP moiety may be required for the interaction of MtRv1421-NTD as a nucleotide-binding protein with its ligand.

Simulation and mechanism for the Ultrasound-Assisted Oiling-Out Process: A case study using Fructose-1,6-diphosphate.

Liquid-liquid separation, commonly referred to as oiling-out, frequently can occurs during crystallization, especially the anti-solvent crystallization process of phosphoryl compounds, and poses potential hurdle for high-quality product. Efficiently regulating oiling-out during crystallization remains a significant challenge. Among various techniques, ultrasound emerges as a green and effective approach to enhance the crystallization process. However, there is a dearth of in-depth research exploring the microscopic mechanisms of this process. Therefore, our research focused on the fructose-1,6-diphosphate (FDP), a typical phosphoryl compound, to gain a deeper understanding of how ultrasound influences the oiling-out process. The focused beam reflectance measurement (FBRM) technology was used to investigate the oiling-out phenomenon of FDPNa3 across various solvent ratios. In addition, the influence of ultrasound on the induction time was studied and the nucleation energy barrier was calculated. Finally, to further unravel the microscopic mechanisms, we utilized molecular simulation techniques to analyze the impact of ultrasound power on the dissolution-precipitation process. Our observations revealed a consistent oiling-out process that attainted a stable state regardless of the solvent employed. Notably, the results of the oiling-out induction time experiments indicated that ultrasound significantly reduced helped lower the nucleation energy barrier of FDP3- ions, thereby dismantling FDP3-clusters in solution. Thus, in turn, shortened the reduced induction time and promoted crystallization. Furthermore, ultrasound reduced the interactions between FDP3-ions and water molecules as well as FDP3- ions themselves. As simulated field intensity increased, these interaction forces gradually diminished, the thickness of the hydration layer surrounding the FDP3- clusters facilitating the disruption of clusters, ultimately enhancing the crystallization process.