1,4-DPCA

In mouse B6 cells, 1,4-DPCA (24 hours) can treat many elevated target gene expressions, such as the pro-angiogenic target genes Vegfa and Hmox1 and the pro-glycolytic target genes Ldh-a, Pgk1, Pdk1, and 1. ZR-75-1 (20 μM 1,4-DPCA) and T4-2 (10 μM 1,4-DPCA) cells showed a significant reduction in colony size following 4-DPCA therapy. In three-dimensional culture, T4-2 cells treated with DPCA produced localized spheroids. Treatment with 1,4-DPCA significantly decreased MDA-MB-231 (10 μM 1,4-DPCA), ZR-75-1, MDA-MB-157, and T4-2 cells [3].

在小鼠 B6 细胞中，1,4-DPCA（24 小时）可以治疗许多升高的靶基因表达，例如促血管生成靶基因 Vegfa 和 Hmox1 以及促糖酵解靶基因 Ldh-a、Pgk1、Pdk1 和 1 ZR-75-1 (20 μM 1,4-DPCA) 和 T4-2 (10 μM 1,4-DPCA) 细胞在 4-DPCA 治疗后显示集落大小显着减小。在三维培养中，用 DPCA 处理的 T4-2 细胞产生了局部球体。用 1,4-DPCA 处理可显着减少 MDA-MB-231 (10 μM 1,4-DPCA)、ZR-75-1、MDA-MB-157 和 T4-2 细胞 [3]。

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用 1,4-DPCA/水凝胶（Gd）处理B6小鼠耳成纤维细胞24小时，通过免疫组织化学和Western blot分析检测到细胞质和细胞核中HIF-1α蛋白表达增加，但不影响HIF-2α表达[1]
对经 1,4-DPCA/水凝胶处理的B6小鼠成纤维细胞的mRNA进行逆转录聚合酶链反应分析，显示多个HIF-1α靶基因表达上调，包括促血管生成基因Vegfa和Hmox1，以及促糖酵解基因Ldh-a、Pgk1、Pdk1和Glut1[1]
用针对Hif1a的siRNA处理可阻断 1,4-DPCA/水凝胶引起的所有上述靶基因的上调[1]
体外用 1,4-DPCA/凝胶处理B6小鼠耳成纤维细胞24小时，细胞对HIF-1α和一系列未成熟细胞标志物呈阳性染色，其染色模式与MRL小鼠耳成纤维细胞无法区分[1]
定量PCR证实，与未处理的细胞相比，经药物/凝胶处理的B6小鼠耳成纤维细胞中干细胞标志物mRNA显著上调[1]
用siHif1a处理NANOG阳性的MRL小鼠耳源性成纤维细胞，导致NANOG蛋白表达几乎完全被抑制[1]

在血管内腹膜聚(锁-共-乙醇酸) (PLGA) 椎间盘中，用 1,4-DPCA 治疗可成功阻止结缔组织向内生长 28 天。 1,4-DPCA 捕获成功地抑制了枢轴椎间盘的内表面和外表面限制性胶原沉积以及结缔组织向内生长，但抑制程度与糖皮质激素不同[2]。

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向瑞士韦伯斯特小鼠皮下注射含有 1,4-DPCA 的水凝胶，在5天内增加了耳组织中HIF-1α蛋白的稳定表达，提供了药物体内释放的功能性测量指标[1]
在10天期间多次皮下注射 1,4-DPCA/水凝胶，导致瑞士韦伯斯特小鼠耳洞穿孔伤后实现完全再生性伤口愈合，愈合持续至第35天[1]
1,4-DPCA/凝胶诱导的耳洞闭合可被同时给予的siHif1a处理所阻断，表明愈合效果依赖于HIF-1α的表达[1]
1,4-DPCA/凝胶处理在瑞士韦伯斯特小鼠中诱导了再生的数个标志：加速再上皮化、细胞去分化、基底膜分解、疤痕形成减少以及后期事件如软骨生成和新毛囊形成[1]
Western blot分析显示，经药物/凝胶处理的瑞士韦伯斯特小鼠在伤后第7、14和21天HIF-1α蛋白表达显著增加，与再生性MRL小鼠中所见水平相似，而HIF-2α表达几乎未受影响[1]

化合物 1,4-DPCA 是脯氨酰羟化酶的有效抑制剂，PHD在常氧条件下介导HIF-1α蛋白的羟基化和后续降解[1]
通过阻断PHD活性，1,4-DPCA 可稳定HIF-1α蛋白[1]

用于体外研究的小鼠耳源性成纤维样细胞进行培养[1]
为测试药物效果，用正常培养基、单独水凝胶或含有 1,4-DPCA 的水凝胶培养细胞。水凝胶前体在孔板中形成固态圆片[1]
处理24小时后，对细胞进行分析。通过免疫染色和Western blot分析评估HIF-1α蛋白表达[1]
对于基因表达分析，从处理的细胞中提取mRNA，通过RT-PCR或定量PCR分析HIF-1α靶基因和干细胞标志物[1]
对于siRNA敲低实验，使用转染试剂将Hif1a siRNA或乱序siRNA转染至70%融合度的细胞中。在转染后48小时检查敲低效率[1]

A locally injectable hydrogel was designed for controlled delivery of 1,4-DPCA over 4 to 10 days. The hydrogel was composed of cross-linked polyethylene glycol molecules (precursors P8Cys and P8NHS) that gel rapidly under physiological conditions[1]
Drug-loaded hydrogels were formed by suspending polymer (Pluronic F127)-stabilized 1,4-DPCA microcrystals in an aqueous mixture of the hydrogel precursors, which solidified rapidly to entrap the drug microcrystals[1]
Swiss Webster mice (8-10 weeks old) received a 2.1 mm through-and-through ear hole punch injury[1]
For single-dose kinetics study: Mice were injected subcutaneously at the back of the neck with 100 µL of hydrogel containing 1,4-DPCA at 2 mg/mL (Gd) or 0 mg/mL (gel alone, G0) several hours after injury. Ear tissue was harvested daily for 5 days for analysis[1]
For regenerative response study: Mice were injected subcutaneously at the back of the neck with 100 µL of Gd or G0 on days 0, 5, and 10 after injury (into three adjacent sites). Ear hole closure was monitored by measuring hole diameters over 35 days[1]
In some experiments, to test distal effects, mice were injected subcutaneously with drug/gel into the left and right rear flank regions on days 0, 5, and 10[1]
For siRNA inhibition in vivo: siHif1a was mixed with an in vivo transfection reagent following the manufacturer's instructions and injected subcutaneously every 48 hours for 20 days beginning on day 0, concurrently with drug/gel treatment[1]
Ear hole diameters were measured in the direction of the ear pinna long axis and the perpendicular axis[1]
At endpoints, mice were euthanized, and ear tissue was collected for histological analysis (e.g., H&E, Alcian blue, picrosirius red staining), immunohistochemistry, Western blot, and RNA extraction for qPCR[1]

[1]. Drug-induced regeneration in adult mice. Sci Transl Med. 2015 Jun 3;7(290):290ra92.

[2]. Transient inhibition of connective tissue infiltration and collagen deposition into porous poly(lactic-co-glycolic acid) discs. J Biomed Mater Res A. 2013 Dec;101(12):3599-606.

[3]. Prolyl-4-hydroxylase α subunit 2 promotes breast cancer progression and metastasis by regulating collagen deposition. BMC Cancer. 2014 Jan 2;14:1.

1,4-DPCA (1,4-dihydrophenonthrolin-4-one-3-carboxylic acid) is a small-molecule inhibitor of prolyl hydroxylases (PHDs)[1]
Hypoxia-inducible factor 1α (HIF-1α) is a central regulator identified in the spontaneous regenerative healing of MRL mice. Its protein is degraded under normoxia via PHD-mediated hydroxylation[1]
Stabilization of HIF-1α via PHD inhibition by 1,4-DPCA is proposed to mimic a key pro-regenerative mechanism found in regenerative MRL mice, promoting processes like aerobic glycolysis, cell migration, inflammation, and stem cell marker expression[1]
Formulating the poorly soluble 1,4-DPCA as microcrystals within a biocompatible hydrogel allowed for sustained local delivery and avoided potential cytotoxicity associated with direct crystal-cell contact[1]
The study demonstrates that pharmacological stabilization of HIF-1α with 1,4-DPCA/hydrogel can induce a regenerative healing response in a normally non-regenerating mammal (Swiss Webster mouse) after injury[1]

C13H8N2O3

240.22

240.053

331830-20-7

459803

White to off-white solid powder

1.5±0.1 g/cm3

474.5±45.0 °C at 760 mmHg

240.8±28.7 °C

0.0±1.2 mmHg at 25°C

1.727

3.28

83.05

2

5

1

18

419

0

XZZHOJONZJQARN-UHFFFAOYSA-N

InChI=1S/C13H8N2O3/c16-12-8-4-3-7-2-1-5-14-10(7)11(8)15-6-9(12)13(17)18/h1-6H,(H,15,16)(H,17,18)

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

配方 1 中的溶解度: ≥ 0.92 mg/mL (3.83 mM) (饱和度未知) in 10% DMSO + 90% Corn Oil (这些助溶剂从左到右依次添加，逐一添加), 澄清溶液。
例如，若需制备1 mL的工作液，可将100 μL 9.2 mg/mL 澄清 DMSO 储备液加入到 900 μL 玉米油中并混合均匀。

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配方 2 中的溶解度: 10 mg/mL (41.63 mM) in 50% PEG300 50% Saline (这些助溶剂从左到右依次添加，逐一添加), 悬浊液; 超声助溶。
*生理盐水的制备：将 0.9 g 氯化钠溶解在 100 mL ddH₂O中，得到澄清溶液。

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请根据您的实验动物和给药方式选择适当的溶解配方/方案：
1、请先配制澄清的储备液(如：用DMSO配置50 或 100 mg/mL母液(储备液));
2、取适量母液，按从左到右的顺序依次添加助溶剂，澄清后再加入下一助溶剂。以 下列配方为例说明 (注意此配方只用于说明，并不一定代表此产品 的实际溶解配方):
10% DMSO → 40% PEG300 → 5% Tween-80 → 45% ddH2O (或 saline);
假设最终工作液的体积为 1 mL, 浓度为5 mg/mL: 取 100 μL 50 mg/mL 的澄清 DMSO 储备液加到 400 μL PEG300 中，混合均匀/澄清；向上述体系中加入50 μL Tween-80，混合均匀/澄清；然后继续加入450 μL ddH2O (或 saline)定容至 1 mL；
3、溶剂前显示的百分比是指该溶剂在最终溶液/工作液中的体积所占比例;
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