Inhibition of glycolysis enhances the efficacy of immunotherapy via PDK-mediated upregulation of PD-L1.

BACKGROUND: Immunotherapy for gastric cancer remains a challenge due to its limited efficacy. Metabolic reprogramming toward glycolysis has emerged as a promising avenue for enhancing the sensitivity of tumors to immunotherapy. Pyruvate dehydrogenase kinases (PDKs) play pivotal roles in regulating glycolysis. The importance of PDKs in the context of gastric cancer immunotherapy and their potential as therapeutic targets have not been fully explored. METHODS: PDK and PD-L1 expression was analyzed using data from the GSE66229 and The Cancer Genome Atlas (TCGA) cohorts. Additionally, the Immune Checkpoint Blockade Therapy Atlas (ICBatlas) database was utilized to assess PDK expression in an immune checkpoint blockade (ICB) therapy group. Subsequently, the upregulation of PD-L1 and the enhancement of anticancer effects achieved by targeting PDK were validated through in vivo and in vitro assays. The impact of PDK on histone acetylation was investigated using ChIP‒qPCR to detect changes in histone acetylation levels. RESULTS: Our analysis revealed a notable negative correlation between PD-L1 and PDK expression. Downregulation of PDK led to a significant increase in PD-L1 expression. PDK inhibition increased histone acetylation levels by promoting acetyl-CoA generation. The augmentation of acetyl-CoA production and concurrent inhibition of histone deacetylation were found to upregulate PD-L1 expression in gastric cancer cells. Additionally, we observed a significant increase in the anticancer effect of PD-L1 antibodies following treatment with a PDK inhibitor. CONCLUSIONS: Downregulation of PDK in gastric cancer cells leads to an increase in PD-L1 expression levels, thus potentially improving the efficacy of PD-L1 immune checkpoint blockade therapy.

Identification of a (+)-cubenene synthase from filamentous fungi Acremonium chrysogenum.

Sesquiterpene synthases convert farnesyl diphosphate into various sesquiterpenes, which find wide applications in the food, cosmetics and pharmaceutical industries. Although numerous putative sesquiterpene synthases have been identified in fungal genomes, many lack biochemical characterization. In this study, we identified a putative terpene synthase AcTPS3 from Acremonium chrysogenum. Through sequence analysis and in vitro enzyme assay, AcTPS3 was identified as a sesquiterpene synthase. To obtain sufficient product for NMR testing, a metabolic engineered Saccharomyces cerevisiae was constructed to overproduce the product of AcTPS3. The major product of AcTPS3 was identified as (+)-cubenene (55.46%) by GC-MS and NMR. Thus, AcTPS3 was confirmed as (+)-cubenene synthase, which is the first report of (+)-cubenene synthase. The optimized S. cerevisiae strain achieved a biosynthesis titer of 597.3 mg/L, the highest reported for (+)-cubenene synthesis.

Hydrogen Sulfide Alleviates Liver Injury Through the S-Sulfhydrated-Kelch-Like ECH-Associated Protein 1/Nuclear Erythroid 2-Related Factor 2/Low-Density Lipoprotein Receptor-Related Protein 1 Pathway.

BACKGROUND AND AIMS: Protein S-sulfhydration mediated by H2 S has been shown to play important roles in several diseases. However, its precise role in liver disease and the related mechanism remain unclear. APPROACH AND RESULTS: We showed that in streptozotocin (STZ)-treated and high-fat diet (HFD)-treated low-density lipoprotein receptor-negative (LDLr-/- ) mice, the H2 S donor GYY4137 ameliorated liver injury, decreased serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) levels, mitigated lipid deposition, and reduced hepatocyte death. Strikingly, S-sulfhydration of Kelch-like ECH-associated protein 1 (Keap1) was decreased in the livers of patients with fatty liver under diabetic conditions. In STZ+HFD-treated LDLr-/- mice and in high glucose-treated and oxidized low-density lipoprotein (ox-LDL)-treated primary mouse hepatocytes, the GYY4137-mediated increase in Keap1 S-sulfhydration induced nuclear erythroid 2-related factor 2 (Nrf2) dissociation from Keap1, which enhanced the nuclear translocation of Nrf2 itself and the consequent expression of antioxidant proteins. Keap1 Cys151 mutation significantly reduced Keap1 S-sulfhydration and abolished the hepatoprotective effects of H2 S both in vivo and in vitro. Nrf2 deficiency inhibited the H2 S-induced beneficial impacts in Nrf2-/- mice. Similarly, in CCl4 -stimulated mice, GYY4137 increased Keap1 S-sulfhydration, improved liver function, alleviated liver fibrosis, decreased hepatic oxidative stress, and activated the Nrf2 signaling pathway; and these effects were abrogated after Keap1 Cys151 mutation. Moreover, H2 S increased the binding of Nrf2 to the promoter region of LDLr-related protein 1 (Lrp1) and consequently up-regulated LRP1 expression, but these effects were disrupted by Keap1 Cys151 mutation. CONCLUSIONS: H2 S-mediated Keap1 S-sulfhydration alleviates liver damage through activation of Nrf2. Hence, administration of exogenous H2 S in the form of the H2 S donor GYY4137 may be of therapeutic benefit in the context of concurrent hyperlipidemia and hyperglycemia-induced or CCl4 -stimulated liver dysfunction.

Identification and characterization of novel MPC1 gene variants causing mitochondrial pyruvate carrier deficiency.

Pyruvate, the end product of glycolysis, is a key metabolic molecule enabling mitochondrial adenosine triphosphate synthesis and takes part in multiple biosynthetic pathways within mitochondria. The mitochondrial pyruvate carrier (MPC) plays a vital role in transporting pyruvate from the cytosol into the organelle. In humans, MPC is a hetero-oligomeric complex formed by the MPC1 and MPC2 paralogs that are both necessary to stabilize each other and form a functional MPC. MPC deficiency (OMIM#614741) due to pathogenic MPC1 variants is a rare autosomal recessive disease involving developmental delay, microcephaly, growth failure, and increased serum lactate and pyruvate. To date, two MPC1 variants in four cases have been reported, though only one with a detailed clinical description. Herein, we report three novel pathogenic MPC1 variants in six patients from three unrelated families, identified within European, Kuwaiti, and Chinese mitochondrial disease patient cohorts, one of whom presented as a Leigh-like syndrome. Functional analysis in primary fibroblasts from the patients revealed decreased expression of MPC1 and MPC2. We rescued pyruvate-driven oxygen consumption rate in patient's fibroblasts by reconstituting with wild-type MPC1. Complementing homozygous MPC1 mutant cDNA with CRISPR-deleted MPC1 C2C12 cells verified the mechanism of variants: unstable MPC complex or ablated pyruvate uptake activity. Furthermore, we showed that glutamine and beta-hydroxybutyrate were alternative substrates to maintain mitochondrial respiration when cells lack pyruvate. In conclusion, we expand the clinical phenotypes and genotypes associated with MPC deficiency, with our studies revealing glutamine as a potential therapy for MPC deficiency.

Identification of the sesquiterpene synthase AcTPS1 and high production of (-)-germacrene D in metabolically engineered Saccharomyces cerevisiae.

BACKGROUND: The sesquiterpene germacrene D is a highly promising product due to its wide variety of insecticidal activities and ability to serve as a precursor for many other sesquiterpenes. Biosynthesis of high value compounds through genome mining for synthases and metabolic engineering of microbial factories, especially Saccharomyces cerevisiae, has been proven to be an effective strategy. However, there have been no studies on the de novo synthesis of germacrene D from carbon sources in microbes. Hence, the construction of the S. cerevisiae cell factory to achieve high production of germacrene D is highly desirable. RESULTS: We identified five putative sesquiterpene synthases (AcTPS1 to AcTPS5) from Acremonium chrysogenum and the major product of AcTPS1 characterized by in vivo, in vitro reaction and NMR detection was revealed to be (-)-germacrene D. After systematically comparing twenty-one germacrene D synthases, AcTPS1 was found to generate the highest amount of (-)-germacrene D and was integrated into the terpene precursor-enhancing yeast strain, achieving 376.2 mg/L of (-)-germacrene D. Iterative engineering was performed to improve the production of (-)-germacrene D, including increasing the copy numbers of AcTPS1, tHMG1 and ERG20, and downregulating or knocking out other inhibitory factors (such as erg9, rox1, dpp1). Finally, the optimal strain LSc81 achieved 1.94 g/L (-)-germacrene D in shake-flask fermentation and 7.9 g/L (-)-germacrene D in a 5-L bioreactor, which is the highest reported (-)-germacrene D titer achieved to date. CONCLUSION: We successfully achieved high production of (-)-germacrene D in S. cerevisiae through terpene synthase mining and metabolic engineering, providing an impressive example of microbial overproduction of high-value compounds.

S-nitrosylation of Hsp90 promotes cardiac hypertrophy in mice through GSK3ß signaling.

Cardiac hypertrophy, as one of the major predisposing factors for chronic heart failure, lacks effective interventions. Exploring the pathogenesis of cardiac hypertrophy will reveal potential therapeutic targets. S-nitrosylation is a kind of posttranslational modification that occurs at active cysteines of proteins to mediate various cellular processes. We here identified heat shock protein 90 (Hsp90) as a highly S-nitrosylated target in the hearts of rodents with hypertrophy, and the role of Hsp90 in cardiac hypertrophy remains undefined. The S-nitrosylation of Hsp90 (SNO-Hsp90) levels were elevated in angiotensin II (Ang II)- or phenylephrine (PE)-treated neonatal rat cardiomyocytes (NRCMs) in vitro as well as in cardiomyocytes isolated from mice subjected to transverse aortic constriction (TAC) in vivo. We demonstrated that the elevated SNO-Hsp90 levels were mediated by decreased S-nitrosoglutathione reductase (GSNOR) expression during cardiac hypertrophy, and delivery of GSNOR adeno-associated virus expression vectors (AAV9-GSNOR) decreased the SNO-Hsp90 levels to attenuate cardiac hypertrophy. Mass spectrometry analysis revealed that cysteine 589 (Cys589) might be the S-nitrosylation site of Hsp90. Delivery of the mutated AAV9-Hsp90-C589A inhibited SNO-Hsp90 levels and attenuated cardiac hypertrophy. We further revealed that SNO-Hsp90 led to increased interaction of glycogen synthase kinase 3ß (GSK3ß) and Hsp90, leading to elevated GSK3ß phosphorylation and decreased eIF2Bε phosphorylation, thereby aggravating cardiac hypertrophy. Application of GSK3ß inhibitor TWS119 abolished the protective effect of Hsp90-C589A mutation in Ang II-treated NRCMs. In conclusion, this study demonstrates a critical role of SNO-Hsp90 in cardiac hypertrophy, which may be of a therapeutic target for cardiac hypertrophy treatment.

Investigation of epidemiological characteristics and development of a nomogram to predict survival in primary ocular adnexal lymphoma.

BACKGROUND: Studies on the epidemiology and prognosis of primary ocular adnexal lymphoma (POAL) are scarce for its low occurrence. The goal of our research was to assess the epidemiologic characteristics, prognostic variables and survival of POAL patients. METHODS: The Surveillance, Epidemiology and End Results (SEER) database was used to collect data on patients identified with POAL from 1975 to 2011 and the incidence rate of POAL from 1975 to 2017. To discover independent predictive markers for overall survival (OS) and disease-specific survival (DSS), multivariable Cox regression analysis was utilised. The independent prognostic factors found by multivariate Cox regression analysis were used to generate a nomogram. RESULTS: A total of 2839 patients were diagnosed with POAL, with an average age of 64.1 years. The total incidence of POAL was 2.51/1000000 (according to the overall adjustment of the 2000 American standard) from 1975 to 2017, and the annual percentage change (APC) was 2.47 (95% confidence interval 1.64-3.32, p < 0.05), showing a sharp upward trend. After multivariate Cox regression analysis, age, gender, year of diagnosis, marital status, primary site, laterality, pathological type and treatment strategy were evaluated as independent prognostic factors of OS or DSS (p < 0.05). A nomogram was constructed to forecast the DSS of 1, 3, 5 and 10 years. The concordance index (C-index) and the calibration plots demonstrated the robustness and accuracy of the nomogram. CONCLUSIONS: Although POAL is sporadic, the incidence has generally increased in the past 36 years. In recent years, survival rates have risen, and radiotherapy can render better OS and DSS. The nomogram specially made for POAL is robust and precise in predicting the DSS of 1, 3, 5 and 10 years.

CRL4 antagonizes SCFFbxo7-mediated turnover of cereblon and BK channel to regulate learning and memory.

Intellectual disability (ID), one of the most common human developmental disorders, can be caused by genetic mutations in Cullin 4B (Cul4B) and cereblon (CRBN). CRBN is a substrate receptor for the Cul4A/B-DDB1 ubiquitin ligase (CRL4) and can target voltage- and calcium-activated BK channel for ER retention. Here we report that ID-associated CRL4CRBN mutations abolish the interaction of the BK channel with CRL4, and redirect the BK channel to the SCFFbxo7 ubiquitin ligase for proteasomal degradation. Glioma cell lines harbouring CRBN mutations record density-dependent decrease of BK currents, which can be restored by blocking Cullin ubiquitin ligase activity. Importantly, mice with neuron-specific deletion of DDB1 or CRBN express reduced BK protein levels in the brain, and exhibit similar impairment in learning and memory, a deficit that can be partially rescued by activating the BK channel. Our results reveal a competitive targeting of the BK channel by two ubiquitin ligases to achieve exquisite control of its stability, and support changes in neuronal excitability as a common pathogenic mechanism underlying CRL4CRBN-associated ID.

Surface display of organophosphorus-degrading enzymes on the recombinant spore of Bacillus subtilis.

Organophosphorus-degrading enzymes show high hydrolysis efficiency and provide an environmentally friendly solution to the pollution of organophosphorus compound. However, poor enzyme stability and tedious purification process have limited practical applications. Spore-based display system can provide many advantages, such as safety, low cost, easy preparation and high resistance to harsh conditions. Recently, we have constituted the recombinant spore displaying organophosphorus hydrolase and organophosphorus acid anhydrolase. In the spore display systems, recombinant spores could be reliably produced and normal sporulation was not affected; the activities of recombinant spores were 15.81 and 10.67 U/mg spores (dry weight) respectively; furthermore, the recombinant spores exhibited significantly enhanced resistance to various harsh conditions compared to free-form enzymes. These results indicated that the spore display could contribute to the practical application of organophosphorus-degrading enzymes and provide a promising solution to bioremediation of organophosphorus compounds.

Bacillus subtilis Spore Surface Display of Haloalkane Dehalogenase DhaA.

The haloalkane dehalogenase DhaA can degrade sulfur mustard (2,2'-dichlorethyl sulfide; also known by its military designation HD) in a rapid and environmentally safe manner. However, DhaA is sensitive to temperature and pH, which limits its applications in natural or harsh environments. Spore surface display technology using resistant spores as a carrier to ensure enzymatic activity can reduce production costs and extend the range of applications of DhaA. To this end, we cloned recombinant Bacillus subtilis spores pHY300PLK-cotg-dhaa-6his/DB104(FH01) for the delivery of DhaA from Rhodococcus rhodochrous NCIMB 13064. A dot blotting showed that the fusion protein CotG-linker-DhaA accounted for 0.41% ± 0.03% (P < 0.01) of total spore coat proteins. Immunofluorescence analyses confirmed that DhaA was displayed on the spore surface. The hydrolyzing activity of DhaA displayed on spores towards the HD analog 2-chloroethyl ethylsulfide was 1.74 ± 0.06 U/mL (P < 0.01), with a specific activity was 0.34 ± 0.04 U/mg (P < 0.01). This is the first demonstration that DhaA displayed on the surface of B. subtilis spores retains enzymatic activity, which suggests that it can be used effectively in real-world applications including bioremediation of contaminated environments.

Activation of c-Abl Kinase Potentiates the Anti-myeloma Drug Lenalidomide by Promoting DDA1 Protein Recruitment to the CRL4 Ubiquitin Ligase.

Cullin-RING ligase 4 (CRL4), a complex of Cul4 and DDB1, regulates the cell cycle, DNA damage repair, and chromatin replication by targeting a variety of substrates for ubiquitination. CRL4 is also hijacked by viral proteins or thalidomide-derived compounds to degrade host restriction factors. Here we report that the c-Abl non-receptor kinase phosphorylates DDB1 at residue Tyr-316 to recruit a small regulatory protein, DDA1, leading to increased substrate ubiquitination. Pharmacological inhibition or genetic ablation of the Abl-DDB1-DDA1 axis decreases the ubiquitination of CRL4 substrates, including IKZF1 and IKZF3, in lenalidomide-treated multiple myeloma cells. Importantly, panobinostat, a recently approved anti-myeloma drug, and dexamethasone enhance lenalidomide-induced substrate degradation and cytotoxicity by activating c-Abl, therefore providing a mechanism underlying their combination with lenalidomide to treat multiple myeloma.

Enhanced adaptive DA-ML carrier phase estimator and its application to accurate laser linewidth and SNR estimation.

We proposed a novel adaptive carrier phase estimator based on the phase information of the received signal only. Through eliminating the perturbation due to the amplitude of the AWGN, the proposed method outperforms the conventional adaptive filter in terms of both the carrier phase estimation and the filter gain tracking. Additionally, a dynamic tracking of both the laser linewidth and SNR is derived based on the proposed carrier phase estimator which requires no prior knowledge of the channel parameters. Numerical simulations and experiments are conducted to verify its feasibility in real applications.

Performance of laser inter-satellite links with dynamic beam waist adjustment.

In this paper, we propose the idea of dynamic beam waist adjustment for laser inter-satellite communications, and study the performance of this dynamic-beam scheme. The beam waist adjustment is based on continuous detection of the instantaneous pointing error angle, which is performed at the transmitter side. Using a square to approximate the circular detector region, we obtain a closed-form expression for calculating the proportion of power that can be collected by the receiver aperture, and derive a simple algebraic solution for the optimum dynamic beam waist. Due to its simple form, the dynamic beam waist value can be computed in real time at the transmitter, and therefore, the adjustment is practically implementable. It is shown that the performance of laser inter-satellite links with dynamic beam waist is better than that with fixed beam waist.

Pancreatic Extragastrointestinal Stromal Tumor: A Case Report.

Gastrointestinal stromal tumors (GISTs) are soft tissue sarcomas that originate from the mesenchymal cells of the gastrointestinal tract. Extra-GISTs (EGISTs) are caused by sites outside the gastrointestinal tract. We reported a case of EGIST of the pancreas in a 51-year-old woman. Enhanced CT scan showed a rounded, slightly hypointense focus in the head of the pancreas and the right pars compacta of the descending duodenum. Routine laboratory and endocrine tests were unremarkable. The patient underwent laparoscopic surgery. The diagnosis of EGIST was confirmed through histopathological and immunohistochemical examination. The tumor was found to be CD117+, CD34+, and DOG+ with a high risk of malignancy. No recurrence was observed during the nine-month postoperative follow-up.

Establishment of an iPSC line (BCHNDi001-A) from a patient with nicotinamide nucleotide repair system deficiency caused by biallelic NAXD mutations.

NAD(P)HX dehydratase (NAXD) gene is one of the key enzymes encoding the nicotinamide nucleotide repair system, reportedly associated with Encephalopathy, progressive, early-onset, with brain edema and/or leukoencephalopathy, 2 (PEBEL2). Here, we generated an induced pluripotent stem cell (iPSC) line from the dermal fibroblasts (HDFs) of a PEBEL2 patient who carried biallelic mutations, c.101_102delTA(p.Thr35Phefs*63) and c.318C > G (p.Ile160Met) in NAXD. These iPSCs showed stable amplification in vitro, expressed pluripotent markers, and differentiated spontaneously into three germ layers, as well as NAXD mutations with normal karyotype.

Mixed reality guided root canal therapy.

Root canal therapy (RCT) is a widely performed procedure in dentistry, with over 25 million individuals undergoing it annually. This procedure is carried out to address inflammation or infection within the root canal system of affected teeth. However, accurately aligning CT scan information with the patient's tooth has posed challenges, leading to errors in tool positioning and potential negative outcomes. To overcome these challenges, a mixed reality application is developed using an optical see-through head-mounted display (OST-HMD). The application incorporates visual cues, an augmented mirror, and dynamically updated multi-view CT slices to address depth perception issues and achieve accurate tooth localization, comprehensive canal exploration, and prevention of perforation during RCT. Through the preliminary experimental assessment, significant improvements in the accuracy of the procedure are observed. Specifically, with the system the accuracy in position was improved from 1.4 to 0.4 mm (more than a 70% gain) using an Optical Tracker (NDI) and from 2.8 to 2.4 mm using an HMD, thereby achieving submillimeter accuracy with NDI. 6 participants were enrolled in the user study. The result of the study suggests that the average displacement on the crown plane of 1.27 ± 0.83 cm, an average depth error of 0.90 ± 0.72 cm and an average angular deviation of 1.83 ± 0.83°. Our error analysis further highlights the impact of HMD spatial localization and head motion on the registration and calibration process. Through seamless integration of CT image information with the patient's tooth, our mixed reality application assists dentists in achieving precise tool placement. This advancement in technology has the potential to elevate the quality of root canal procedures, ensuring better accuracy and enhancing overall treatment outcomes.

Long-term effectiveness and tolerability of ketogenic diet therapy in patients with genetic developmental and epileptic encephalopathy onset within the first 6 months of life.

OBJECTIVE: To investigate the effectiveness and tolerability of ketogenic diet therapy (KDT) in patients with developmental and epileptic encephalopathy (DEE) associated with genetic etiology which onset within the first 6 months of life, and to explore the association between response to KDT and genotype/clinical parameters. METHODS: We retrospectively reviewed data from patients with genetic DEE who started KDT at Beijing Children's Hospital between January 1, 2016, and December 31, 2021. RESULTS: A total of 32 patients were included, involving 14 pathogenic or likely pathogenic single genes, and 16 (50.0%) patients had sodium/potassium channel gene variants. The median age at onset of epilepsy was 1.0 (IQR: 0.1, 3.0) months. The median age at initiation of KDT was 10.0 (IQR: 5.3, 13.8) months and the median duration of maintenance was 14.0 (IQR: 7.0, 26.5) months, with a mean blood ß-hydroxybutyrate of 2.49 ± 0.62 mmol/L. During the maintenance period of KDT, 26 (81.3%) patients had a ≥50% reduction of seizure frequency, of which 12 (37.5%) patients achieved seizure freedom. Better responses were observed in patients with STXBP1 variants, with four out of five patients achieving seizure freedom. There were no statistically differences in the age of onset, duration of epilepsy before KDT, blood ketone values, or the presence of ion channel gene variants between the seizure-free patients and the others. The most common adverse effects were gastrointestinal side effects, which occurred in 21 patients (65.6%), but all were mild and easily corrected. Only one patient discontinued KDT due to nephrolithiasis. SIGNIFICANCE: KDT is effective in treating early onset genetic DEE, and no statistically significant relationship has been found between genotype and effectiveness in this study. KDT is well tolerated in most young patients, with mild and reversible gastrointestinal side effects being the most common, but usually not the reason to discontinue KDT. PLAIN LANGUAGE SUMMARY: This study evaluated the response and side effects of ketogenic diet therapy (KDT) in patients who had seizures within the first 6 months of life, and were diagnosed with genetic developmental and epileptic encephalopathy (DEE), a type of severe epilepsy with developmental delay caused by gene variants. Thirty-two patients involving 14 gene variants who started KDT at Beijing Children's Hospital between were included. KDT was effective in treating early onset genetic DEE in this cohort, and patients with STXBP1 variants responded better; however, no statistically significant relationship was found between gene variant and response. Most young patients tolerated KDT well, with mild and reversible gastrointestinal side effects being the most common.

Development of a novel fluorescent protein-based probe for efficient detection of Pb2+ in serum inspired by the metalloregulatory protein PbrR691.

BACKGROUND: The accurate and rapid detection of blood lead concentration is of paramount importance for assessing human lead exposure levels. Fluorescent protein-based probes, known for their high detection capabilities and low toxicity, are extensively used in analytical sciences. However, there is currently a shortage of such probes designed for ultrasensitive detection of Pb2+, and no reported probes exist for the quantitative detection of Pb2+ in blood samples. This study aims to fill this critical void by developing and evaluating a novel fluorescent protein-based probe that promises accurate and rapid lead quantification in blood. RESULTS: A simple and small-molecule fluorescent protein-based probe was successfully constructed herein using a peptide PbrBD designed for Pb2+ recognition coupled to a single fluorescent protein, sfGFP. The probe retains a three-coordinate configuration to identify Pb2+ and has a high affinity for it with a Kd' of 1.48 ± 0.05 × 10-17 M. It effectively transfers the conformational changes of the peptide to the chromophore upon Pb2+ binding, leading to fast fluorescence quenching and a sensitive response to Pb2+. The probe offers a broad dynamic response range of approximately 37-fold and a linear detection range from 0.25 nM to 3500 nM. More importantly, the probe can resist interference of metal ions in living organisms, enabling quantitative analysis of Pb2+ in the picomolar to millimolar range in serum samples with a recovery percentage of 96.64%-108.74 %. SIGNIFICANCE: This innovative probe, the first to employ a single fluorescent protein-based probe for ultrasensitive and precise analysis of Pb2+ in animal and human serum, heralds a significant advancement in environmental monitoring and public health surveillance. Furthermore, as a genetically encoded fluorescent probe, this probe also holds potential for the in vivo localization and concentration monitoring of Pb2+.

Epigenetic regulation of CD38/CD48 by KDM6A mediates NK cell response in multiple myeloma.

Anti-CD38 monoclonal antibodies like Daratumumab (Dara) are effective in multiple myeloma (MM); however, drug resistance ultimately occurs and the mechanisms behind this are poorly understood. Here, we identify, via two in vitro genome-wide CRISPR screens probing Daratumumab resistance, KDM6A as an important regulator of sensitivity to Daratumumab-mediated antibody-dependent cellular cytotoxicity (ADCC). Loss of KDM6A leads to increased levels of H3K27me3 on the promoter of CD38, resulting in a marked downregulation in CD38 expression, which may cause resistance to Daratumumab-mediated ADCC. Re-introducing CD38 does not reverse Daratumumab-mediated ADCC fully, which suggests that additional KDM6A targets, including CD48 which is also downregulated upon KDM6A loss, contribute to Daratumumab-mediated ADCC. Inhibition of H3K27me3 with an EZH2 inhibitor resulted in CD38 and CD48 upregulation and restored sensitivity to Daratumumab. These findings suggest KDM6A loss as a mechanism of Daratumumab resistance and lay down the proof of principle for the therapeutic application of EZH2 inhibitors, one of which is already FDA-approved, in improving MM responsiveness to Daratumumab.

SLC44A2 regulates vascular smooth muscle cell phenotypic switching and aortic aneurysm.

Aortic aneurysm is a life-threatening disease with limited interventions, closely related to vascular smooth muscle cells (VSMCs) phenotypic switching. SLC44A2, a member of solute carrier series 44 (SLC44) family, remains under-characterized in the context of cardiovascular diseases. Venn diagram analysis based on microarray and single-cell RNA sequencing identified SLC44A2 as a major regulator of VSMCs phenotypic switching in aortic aneurysm. Screening for Slc44a2 amongst aortic cell lineages demonstrated its predominant location in VSMCs. Elevated levels of SLC44A2 were evidenced in the aorta of both abdominal aortic aneurysm patients and angiotensin II (Ang II)-infused Apoe-/- mice. In vitro, SLC44A2 silencing promoted VSMCs towards a synthetic phenotype, while SLC44A2 overexpression attenuated VSMCs phenotypic switching. VSMCs-specific SLC44A2 knockout mice were more susceptible to aortic aneurysm under Ang II infusion, while SLC44A2 overexpression showed protective effects. Mechanistically, SLC44A2 interaction with NRP1 and ITGB3 activates TGF-ß/SMAD signaling, thereby promoting contractile genes expression. Elevated SLC44A2 in aortic aneurysm is associated with upregulated runt-related transcription factor 1 (RUNX1). Furthermore, low dose of lenalidomide (LEN) suppressed aortic aneurysm progression by enhancing SLC44A2 expression. These findings reveal SLC44A2/NRP1/ITGB3 complex is a major regulator of VSMCs phenotypic switching and provide potential therapeutic approach (LEN) for aortic aneurysm treatment.