UNASSIGNED: Intravitreal injection of anti-VEGF antibodies remains the primary therapy for exudative age-related macular degeneration (exAMD), although its efficacy is limited. Previous research has demonstrated that both a loss-of-function mutation of srr and the intravenous injection of a serine racemase inhibitor, L-aspartic acid β-hydroxamate (L-ABH), significantly inhibit laser-induced choroidal neovascularization (CNV) in mice. Given that L-ABH is a small molecule, this study investigated the effects of L-ABH administered via eye drops on CNV, aiming to develop a noninvasive treatment strategy for exAMD.
UNASSIGNED: CNV models in mice and rhesus macaques were established through laser photocoagulation. Seven monkeys were randomly assigned to receive either saline solution or L-ABH eye drops. Intraperitoneal or intravenous injection of fluorescein characterized CNV in both mice and monkeys. Fluorescein fundus angiography was used to assess leakage, whereas optical coherence tomography measured retinal thickness in the monkeys.
UNASSIGNED: L-ABH eye drops significantly reduced fluorescein leakage in laser-injured mice (P < 0.001 compared to saline). In laser-injured rhesus macaques, the average percent changes in leakage areas treated with L-ABH were 2.5% ± 25.8% (P = 0.004) and 1.5% ± 75.7% (P = 0.023 compared to saline solution) on day 14 and day 28, respectively. However, L-ABH eye drops did not significantly affect the number of grade IV laser spots or retinal thickness, whereas bevacizumab did.
UNASSIGNED: This study demonstrates the potential efficacy of an SRR inhibitor in two animal models of laser-induced CNV.
UNASSIGNED: This represents the first investigation into the effects of topical delivery of an SRR inhibitor on CNV.

目的： 探讨管状囊性肾细胞癌（tubulocystic renal cell carcinoma，TCRCC）的临床病理学特征。 方法： 回顾性分析新乡医学院第一附属医院2例、会诊1例TCRCC临床病理学资料并进行随访及文献复习。 结果： 例1、2为女性，例3为男性，年龄分别为73、52、47岁。大体检查肿瘤与正常组织边界清楚；镜下肿瘤呈大小不等的囊腔和小到中等的小管样结构，被覆单层肿瘤细胞，细胞界限不清，呈扁平、立方、柱状或鞋钉样，细胞显著增大，胞质嗜酸，核仁明显，核分裂象不易见，小管或囊腔之间为薄层纤维性间质，其中例1间质可见钙盐沉积伴骨化生同时合并透明细胞肾细胞癌［国际泌尿病理协会（ISUP）1级］。免疫表型：3例TCRCC肿瘤细胞细胞角蛋白（CK）19、PAX8、P504s、波形蛋白和FH均阳性，CK7、TFE3均阴性，肾细胞癌标志物（RCC）、CD10、PAX2和34βE12部分病例表达。分子遗传学特征：例1显示第9号染色体丢失，例2显示第9号和17号染色体获得，例3显示第7号染色体获得。随访时间8~24个月，均无病生存。 结论： TCRCC是一种少见的肾细胞癌亚型，病理诊断和鉴别诊断主要依赖于形态学特征，以免疫表型为辅助，分子遗传学检测结果未发现一致性改变，具有惰性的生物学行为，很少复发或转移，可以通过部分或根治性肾切除术治疗。.

Objective: To study the clinicopathological features, immunohistochemical phenotypes, molecular changes, differential diagnosis and prognosis of isolated intraductal carcinoma of the prostate (iIDC-P). Methods: Three iIDC-P cases were collected retrospectively from 2016 to 2022 at Ningbo Clinical Pathology Diagnosis Center, Ningbo, China. The clinicopathologic features and immunophenotypic profiles were studied using light microscopy and immunohistochemistry. A targeted next-generation sequencing panel was used to analyze cancer-associated mutations. Follow-up and literature review were also performed. Results: The patients\' ages were 61, 67 and 77 years, and their preoperative prostate specific antigen (PSA) levels were 7.99, 7.99 and 4.86 μg/L, respectively. Case 1 and 2 were diagnosed on needle biopsy and radical prostatectomy (RP) specimens, and case 3 was diagnosed on a specimen of transurethral resection of the prostate (TURP). The RP specimen was entirely submitted for histologic examination. In the case 1, iIDC-P was found in one tissue core (involving two ducts) in the biopsy specimen, and in 6 sections (diameter, 0.3-1.1 cm) from the radical prostatectomy specimen, and one section had separate foci of low-grade acinar adenocarcinoma (diameter, 0.05 cm). In the case 2, 6 tissue sections from the biopsy specimens showed iIDC-P, and 13 sections from RP specimen showed iIDC-P (diameter, 0.5-1.6 cm), and the other 3 sections had separate low grade acinar adenocarcinoma (diameter, 0.6 cm). In the case 3, 5 tissue blocks from the TURP specimen showed iIDC-P. The case 1 and 2 showed solid architecture with expansile proliferation of neoplastic cells in native ducts and acini. The case 3 showed dense or loose cribriform pattern, with marked cytological atypia, and frequent mitotic figures. Comedonecrosis was found in solid or dense cribriform glands in the case 2. Immunohistochemically, surrounding basal cells were highlighted using high-molecular-weight cytokeratin (34βE12 and CK5/6) and p63, while P504s was positive in the tumor cells. The tumor cells were also positive for AR and prostate markers (NKX3.1, PSA and PSAP), and negative for GATA3. The iIDC-P and acinar adenocarcinoma both showed weak PTEN expression and no ERG (nuclear) expression. In case 2 and 3, targeted sequencing revealed activated oncogenic driver mutations in MAPK and PI3K pathway genes (KRAS, MTOR and PTEN). In addition, pathogenic mutation in TP53 and FOXA1 mutation were found in the case 2 and 3, respectively. No case demonstrated TMPRSS2::ERG translocation. All cases were microsatellite stable and had lower tumor mutation burdens (range, 2.1-3.1 muts/Mb). The patients showed no biochemical recurrence or metastasis after follow-up of 16-91 months. Conclusions: iIDC-P is a special type of intraductal carcinoma of the prostate and differs from intraductal carcinoma within high-grade prostate cancer. iIDC-P has unique molecular characteristics and may represent as a molecularly unique in situ tumor of prostate cancer.
目的： 探讨前列腺孤立性导管内癌（isolated intraductal carcinoma of the prostate，iIDC-P）的临床病理特征、分子改变、鉴别诊断及预后。 方法： 回顾性收集宁波市临床病理诊断中心2016—2022年间的3例iIDC-P的临床病理学资料，进行光镜观察、免疫组织化学染色、高通量DNA靶向测序及随访，并复习相关文献。 结果： 例1~3患者年龄分别为61、67、77岁，术前总前列腺特异性抗原（TPSA）水平分别为7.99、7.99、4.86 μg/L。例1和例2包含前列腺穿刺和根治标本，例3为经尿道前列腺电切标本。镜下：例1穿刺标本仅1条组织见导管内癌（累及2个导管）、根治标本其中6张切片见导管内癌（病灶直径0.3~1.1 cm），其中1张切片见小灶低级别腺泡腺癌（病灶直径0.05 cm）。例2穿刺标本6条组织见导管内癌，根治标本其中13张切片见导管内癌（病灶直径0.5~1.6 cm），另外3张切片可见低级别腺泡腺癌（最大病灶直径0.6 cm）。例3电切标本其中5条组织见导管内癌。例1和例2导管内癌呈实性生长，导管-腺泡显著膨胀，核显著异型；例3呈致密筛状或疏松筛状结构，核显著异型，核仁明显。例2伴有粉刺样坏死，3例核分裂象均易见。导管内癌免疫表型：双标34βE12+P504s显示基底细胞表达34βE12/异型肿瘤细胞表达P540s，p63和CK5/6显示腺泡/导管周围基底细胞，肿瘤细胞表达NKX3.1、雄激素受体、前列腺特异性抗原、前列腺特异性酸性磷酸酶，不表达GATA3，导管内癌和腺泡腺癌均弱表达PTEN、不表达ERG。高通量DNA靶向测序：例2和例3伴有MAPK/PI3K通路基因改变（KRAS、MTOR和PTEN），例2伴有p53突变、例3伴有FOXA1突变。3例均未发现TMPRSS2：：ERG融合，3例均显示微卫星稳定，肿瘤突变负荷低（2.1~3.1 muts/Mb）。随访16~91个月，均无生化复发及远处转移。 结论： iIDC-P是一种特殊的前列腺导管内癌，不同于伴有高级别前列腺癌的前列腺导管内癌，具有独特的分子改变，可能代表一种特殊类型前列腺癌的原位病变。.

BACKGROUND: D-psicose 3-epimerase (DPEase) is a potential catalytic enzyme for D-psicose production. D-psicose, also known as D-allulose, is a low-calorie sweetener that has gained considerable attention as a healthy alternative sweetener due to its notable physicochemical properties. This research focused on an in-depth investigation of the expression of the constructed DPEase gene from Agrobacterium tumefaciens in Escherichia coli for D-psicose synthesis. Experimentally, this research created the recombinant enzyme, explored the optimization of gene expression systems and protein purification strategies, investigated the enzymatic characterization, and then optimized the D-psicose production. Finally, the produced D-psicose syrup underwent acute toxicity evaluation to provide scientific evidence supporting its safety.
RESULTS: The optimization of DPEase expression involved the utilization of Mn2+ as a cofactor, fine-tuning isopropyl β-D-1-thiogalactopyranoside induction, and controlling the induction temperature. The purification process was strategically designed by a nickel column and an elution buffer containing 200 mM imidazole, resulting in purified DPEase with a notable 21.03-fold increase in specific activity compared to the crude extract. The optimum D-psicose conversion conditions were at pH 7.5 and 55 °C with a final concentration of 10 mM Mn2+ addition using purified DPEase to achieve the highest D-psicose concentration of 5.60% (w/v) using 25% (w/v) of fructose concentration with a conversion rate of 22.42%. Kinetic parameters of the purified DPEase were Vmax and Km values of 28.01 mM/min and 110 mM, respectively, which demonstrated the high substrate affinity and efficiency of DPEase conversion by the binding site of the fructose-DPEase-Mn2+ structure. Strategies for maintaining stability of DPEase activity were glycerol addition and storage at -20 °C. Based on the results from the acute toxicity study, there was no toxicity to rats, supporting the safety of the mixed D-fructose-D-psicose syrup produced using recombinant DPEase.
CONCLUSIONS: These findings have direct and practical implications for the industrial-scale production of D-psicose, a valuable rare sugar with a broad range of applications in the food and pharmaceutical industries. This research should advance the understanding of DPEase biocatalysis and offers a roadmap for the successful scale-up production of rare sugars, opening new avenues for their utilization in various industrial processes.

Aging is associated with a decrease in N-methyl-D-aspartate (NMDA) receptor function, which is critical for maintaining synaptic plasticity, learning, and memory. Activation of the NMDA receptor requires binding of the neurotransmitter glutamate and also the presence of co-agonist D-serine at the glycine site. The enzymatic conversion of L-serine to D-serine is facilitated by the enzyme serine racemase (SR). Subsequently, SR plays a pivotal role in regulating NMDA receptor activity, thereby impacting synaptic plasticity and memory processes in the central nervous system. As such, age-related changes in the expression of SR could contribute to decreased NMDA receptor function. However, age-associated changes in SR expression levels in the medial and lateral prefrontal cortex (mPFC, lPFC), and in the dorsal hippocampal subfields, CA1, CA3, and dentate gyrus (DG), have not been thoroughly elucidated. Therefore, the current studies were designed to determine the SR expression profile, including protein levels and mRNA, for these regions in aged and young male and female Fischer-344 rats. Our results demonstrate a significant reduction in SR expression levels in the mPFC and all hippocampal subfields of aged rats compared to young rats. No sex differences were observed in the expression of SR. These findings suggest that the decrease in SR levels may play a role in the age-associated reduction of NMDA receptor function in brain regions crucial for cognitive function and synaptic plasticity.

The sulfite-reducing bacterium Bilophila wadsworthia, a common human intestinal pathobiont, is unique in its ability to metabolize a wide variety of sulfonates to generate sulfite as a terminal electron acceptor (TEA). The resulting formation of H2S is implicated in inflammation and colon cancer. l-cysteate, an oxidation product of l-cysteine, is among the sulfonates metabolized by B. wadsworthia, although the enzymes involved remain unknown. Here we report a pathway for l-cysteate dissimilation in B. wadsworthia RZATAU, involving isomerization of l-cysteate to d-cysteate by a cysteate racemase (BwCuyB), followed by cleavage into pyruvate, ammonia and sulfite by a d-cysteate sulfo-lyase (BwCuyA). The strong selectivity of BwCuyA for d-cysteate over l-cysteate was rationalized by protein structural modeling. A homolog of BwCuyA in the marine bacterium Silicibacter pomeroyi (SpCuyA) was previously reported to be a l-cysteate sulfo-lyase, but our experiments confirm that SpCuyA too displays a strong selectivity for d-cysteate. Growth of B. wadsworthia with cysteate as the electron acceptor is accompanied by production of H2S and induction of BwCuyA. Close homologs of BwCuyA and BwCuyB are present in diverse bacteria, including many sulfate- and sulfite-reducing bacteria, suggesting their involvement in cysteate degradation in different biological environments.

The obligate intracellular bacterium, Chlamydia trachomatis, has evolved to depend on its human host for many metabolites, including most amino acids and three of the four nucleotides. Given this, it is not surprising that depletion of a single amino acid in the host cell growth medium blocks chlamydial replication. Paradoxically, supra-normal levels of some amino acids also block productive replication of Chlamydia. Here, we have determined how elevated serine levels, generated by exogenous supplementation, impede chlamydial inclusion development and reduce the generation of infectious progeny. Our findings reveal that human serine racemase, which is broadly expressed in multiple tissues, potentiates the anti-chlamydial effect of elevated serine concentrations. In addition to reversibly converting l-serine to d-serine, serine racemase also deaminates serine via β-elimination. We have determined that d-serine does not directly impact Chlamydia; rather, ammonia generated by serine deamination limits the productive chlamydial replication. Our findings imply that ammonia produced within host cells can traverse the chlamydial inclusion membrane. Further, this property of serine deaminase can be exploited to sensitize Chlamydia to concentrations of doxycycline that are otherwise not bactericidal. Because exogenously elevated levels of serine can be tolerated over extended periods, the broad expression pattern of serine racemase indicates it to be a host enzyme whose activity can be directed against multiple intracellular bacterial pathogens. From a therapeutic perspective, demonstrating host metabolism can be skewed to generate an anti-bacterial metabolite that synergizes with antibiotics, we believe our results provide a new approach to target intracellular pathogens.

Cancer stands as a prominent global cause of mortality, necessitating early detection to augment survival rates and alleviate economic burdens on healthcare systems. In particular, prostate cancer (PCa), impacting 1.41 million men globally in 2020, accentuates the demand for sensitive and cost-effective detection methods beyond traditional prostate-specific antigen (PSA) testing. While clinical techniques exhibit limitations, biosensors emerge as compact, user-friendly alternatives to traditional laboratory approaches. However, existing biosensors predominantly concentrate on PSA detection, prompting the necessity for advancing toward multiplex sensing platforms. This study introduces a compact opto-microfluidic sensor featuring a substrate of gold nanospikes, fabricated via electrodeposition, for enhanced sensitivity. Embedded within a microfluidic chip, this nanomaterial enables the precise and concurrent measurement of PSA, alongside two complementary PCa biomarkers, matrix metalloproteinase-2 (MMP-2) and anti-α-methylacyl-CoA racemase (anti-AMACR) in diluted human plasma, offering a comprehensive approach to PSA analysis. Taking advantage of the localized surface plasmon resonance principle, this biosensor offers robustness and sensitivity in real sample analysis without the need for labeling agents. With the limit of detection at 0.22, 0.37, and 0.18 ng/mL for PSA, MMP-2, and anti-AMACR, respectively, this biosensing platform holds promise for point-of-care analysis, underscoring its potential impact on medical diagnostics.

D-Allulose 3-epimerase (DAE) is a vital biocatalyst for the industrial synthesis of D-allulose, an ultra-low calorie rare sugar. However, limited thermostability of DAEs hinders their use at high-temperature production. In this research, hyperthermophilic TI-DAE (Tm = 98.4 ± 0.7 ℃) from Thermotoga sp. was identified via in silico screening. A comparative study of the structure and function of site-directed saturation mutagenesis mutants pinpointed the residue I100 as pivotal in maintaining the high-temperature activity and thermostability of TI-DAE. Employing TI-DAE as a biocatalyst, D-allulose was produced from D-fructose with a conversion rate of 32.5%. Moreover, TI-DAE demonstrated excellent catalytic synergy with glucose isomerase CAGI, enabling the one-step conversion of D-glucose to D-allulose with a conversion rate of 21.6%. This study offers a promising resource for the enzyme engineering of DAEs and a high-performance biocatalyst for industrial D-allulose production.

D-allulose is a naturally occurring rare sugar and beneficial to human health. However, the efficient biosynthesis of D-allulose remains a challenge. Here, we mined a new D-tagatose 3-epimerase from Kroppenstedtia eburnean (KeDt3e) with high catalytic efficiency. Initially, crucial factors contributing to the low conversion of KeDt3e were identified through crystal structure analysis, density functional theory calculations (DFT), and molecular dynamics (MD) simulations. Subsequently, based on the mechanism, combining restructuring the flexible region, proline substitution based onconsensus sequence analysis, introducing disulfide bonds, and grafting properties, and reshaping the active center, the optimal mutant M5 of KeDt3e was obtained with enhanced thermostability and activity. The optimal mutant M5 exhibited an enzyme activity of 130.8 U/mg, representing a 1.2-fold increase; Tm value increased from 52.7 °C to 71.2 °C; and half-life at 55 °C extended to 273.7 min, representing a 58.2-fold improvement, and the detailed mechanism of performance improvement was analyzed. Finally, by screening the ribosome-binding site (RBS) of the optimal mutant M5 recombinant bacterium (G01), the engineered strain G08 with higher expression levels was obtained. The engineered strain G08 catalyzed 500 g/L D-fructose to produce 172.4 g/L D-allulose, with a conversion of 34.4% in 0.5 h and productivity of 344.8 g/L/h on a 1 L scale. This study presents a promising approach for industrial-scale production of D-allulose.