HC与ASGP-R在非内因子途径维生素B12肠内吸收中的作用及其机制

Intrinsic factor (IF), a glycoprotein secreted by the parietal cell located at gastric mucosa, is a dispensable factor for the enteral absorption of vitamin B12. Obviously, the patients after total gastrectomy are unable to secrete IF any more, making it impossible to bind and absorb vitamin B12 through IF-B12 complex pattern. Finally, these patients would inevitably suffer from vitamin B12 deficiency. Consequently, intramuscular injection of vitamin B12 is the only known and recommended treatment for vitamin B12 deficiency. However, recently it was suggested that vitamin B12 deficiency patients could also be corrected by consistently taking relatively large doses of vitamin B12 orally, for the patients without IF secretion. This updated knowledge suggested that an enteral non-IF-related absorption mechanism of vitamin B12 must be existed. To our best knowledge, however, the study about this new mechanism is rare. .Haptocorrin (HC), which is secreted by sialic acid gland, is able to mediate the uptake of vitamin b12 through ASGP-R, which is largely expressed in the liver cell membrane. It was confirmed by our updated experiments that ASGP-R can be found in human intestine tract. More importantly, HC and R1 which is a subtype of ASGP-R, are both found to be upregulated significantly, and R2 which is another subtype of ASGP-R to be downregulated among totally gastrectomized patients, compared to those among healthy people. Based on our findings above, we hypothesized that the expression of HC and subtype R1 of ASGP-R might be upregulated, while the expression of subtype R2 of ASGP-R be downregulated in the totally gastrectomized patients, which could be the new mechanism of enteral non-IF-related absorption of vitamin B12. .In the present study, we will create both totally gastrectomized model of rat and human as our objects, aiming to determine the enteral non-IF-related absorption site of vitamin B12, to explore the function of haptocorrin and ASGP-R in this new mechanism, and then to provide a promising strategy for the management of the patients with vitamin B12 deficiency.

胃分泌的内因子（IF）是维生素B12肠内吸收的必需因子。全胃切除后失去IF参与而发生B12缺乏，仅能通过肠外途径肌注B12纠正。近年来研究表明：全胃切除术后B12缺乏亦可通过口服大剂量B12纠正，提示非内因子途径B12肠内吸收途径的存在，但其机制尚不明确。唾液中的结合卟啉蛋白（HC）能与B12形成复合物，被肝细胞膜富集的去唾液酸糖蛋白受体（ASGP-R）识别并吸收。我们的预实验发现，相比正常人，胃切除后患者HC表达上调，小肠黏膜ASGP-R的亚型R1上调、亚型R2下调。因此我们推测，人体缺乏IF时，通过激活HC、ASGP-R亚型R1的上调、亚型R2的下调，介导B12的肠内吸收。本研究以全胃切除大鼠及人体为对象：（1）探索B12肠内吸收部位；（2）探索ASGP-R与HC在B12肠内吸收中的作用及机制。从而阐明非内因子途径维生素B12肠内吸收新机制，为B12缺乏的防治提供新思路。

背景.维生素B12 (VB12) 是一种人体必需的营养元素，口服摄取的VB12在食管内先与结合蛋白（HC）形成HC-VB12复合物后，再与胃壁细胞分泌的内因子（IF）结合，形成IF-VB12复合物运送至回肠末端方能吸收，而在全胃切除缺乏IF机体， HC-VB12复合物难以完成肠内吸收，会导致神经炎、恶性贫血等不可逆性VB12缺乏症。本人前期研究提出：全胃切除的人体内可能存在非内因子途径维生素B12肠内吸收机制。与此同时，胃癌病灶对VB12等营养物质摄取呈显著增加，其相关受体表达显著上调，这提示VB12具有潜在肿瘤靶向性。但目前VB12的简便检测方法难以实现。本研究探索VB12荧光标记探针，在可视化条件下验证VB12肠内吸收、肿瘤摄取机制，最终利用VB12特异吸收通路实现药物的肠内高效吸收与肿瘤靶向给药。..研究内容及关键结果.1.聚集诱导发光（AIE）生物材料具有背景荧光低、生物相容性好、光稳定性好等优势，本人基于AIE效能研发VB12-AIE的可视化荧光探针。该探针可被肠上皮细胞高效摄取，这提示VB12-AIE探针可作为其吸收代谢的监测新手段。.2.前期研究的ASGP-R蛋白未能显著增加HC-VB12的吸收效。本研究进一步利用基因芯片检测胃切除术前/术后肠黏膜蛋白变化，结果显示：促VB12 外排的ABCC1 蛋白表达下调，而PRSS38 蛋白表达上调。利用IF缺乏的肠上皮细胞体外模型，上调细胞内PRSS38 蛋白，可增加HC-VB12的吸收效率，这提示PRSS38可能作为HC肠内结合新靶点，促进VB12的肠内吸收。.3.在消化道重建的胃癌术后人群，传统口服药物吸收效能下降，本人研发VB12结合型海藻酸纳米复合物，实现药物高效肠内吸收。.4.传统药物难以主动到达胃癌病灶，本人研发VB12结合型丝胶蛋白纳米复合物，可通过CD320受体介导的胞吞途径实现药物高效靶向抗肿瘤作用。..研究意义.1.本研究研发了VB12-AIE荧光探针，发现PRSS38蛋白可能作为HC的肠上皮结合蛋白，促进VB12肠内吸收，为VB12吸收代谢、为VB12缺乏人群的营养干预提供理论基础。.2.本研究针对胃癌切除人群合成VB12结合型的纳米复合物，可突破药物口服效能低下、缺乏肿瘤靶向等两大临床难题，在实现精准靶向给药上有着广泛临床应用前景。

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