糖尿病視網膜病變的臨床治療與病理研究

Clinical Treatments and Pathological Studies of Diabetic Retinopathy

葉伯廷(Po-Ting Yeh)

指導教授 ： 林中梧

國立臺灣大學/醫學院/病理學研究所/博士(2018年)

https://doi.org/10.6342/NTU201801377

摘要

玻璃體切除手術目前是增殖性糖尿病視網膜病變各種併發症的主要治療方法之一。以使用現代的手術儀器和技術，手術成功率相當高，從70％到90％不等。成功的關鍵取決於糖尿病視網膜病變疾病發展過程的長短與疾病嚴重程度。手術後，約三分之一手術成功的患者會發生復發性玻璃體出血。出血的原因可能來自殘留的纖維血管組織或是手術後在視網膜周邊形成的新生血管破裂有關。復發性玻璃體出血不僅會延緩病人視力的恢復，還可能會導致需要再次接受手術。因此，若在復雜的糖尿病性玻璃體切除手術中，進行適當的預防性處理程序，以減少術中和術後視網膜出血，這對於預防復發性玻璃體出血是非常重要的。如果術後不幸發生了復發性玻璃體出血，適當的處理可以使新生血管消退，並減少反覆出血的可能性；從而縮短再吸收時間並減少手術需求。在我們的系列研究中，我們設計了一些臨床研究並發展了一些因應不同狀況病人，而有不同處理的有效手術方法，包括(1)術前1週進行玻璃體內bevacizumab（1.25 mg）注射及合併初次矽油灌注；(2)合併玻璃體內bevacizumab注射（1.25 mg）以及視網膜前部冷凍療法來治療糖尿病玻璃體切除術後性玻璃體再次出血；(3)玻璃體內重複注射bevacizumab（1.25 mg）以治療糖尿病視網膜病變經玻璃體切除術後再次出血的患者。我們的結果顯示這些方法對於預防與治療復發性糖尿病玻璃體出血是有效和安全的。
在增殖性糖尿病視網膜病變中，視網膜的纖維血管增生(fibrovascular proliferation)是其獨特的臨床變化；病理上，由於纖維血管增生其複雜的玻璃體與視網膜之間交互關係，頻繁出現的視網膜前膜(epiretinal membrane)和強大的促發炎(proinflammatory)and和促血管生成(pro-angiogenic)環境，進而導致視網膜的病變。視網膜前膜的形成可能發生在各種臨床環境中。它是由視網膜表面的細胞增殖引起的，但是這些增殖細胞的起源目前還是有些爭論。過去，視網膜前膜手術標本中已發現主要有視網膜內膠質細胞(glial cells)，其他有視網膜結締組織細胞，甚至也有視網膜色素上皮細胞(retinal pigment epithelial cells)。視網膜前膜形成的病理生理機制在不同疾病情況下可能變化很大。在糖尿病視網膜病變中，視網膜前膜可能在手術前發生，或在成功玻璃體切除術後發生。玻璃體切除術後眼部的特殊狀況，如局部缺血，炎症，出血和纖維血管膜殘留等，都可能使眼睛容易形成術後的視網膜前膜。為了更好地了解視網膜前膜對經過玻璃體切除手術後形成的原因及視網膜前膜移除後對黃斑部結構和預後的影響，我們進行了一項回顧性研究，以評估可能的危險因素，視網膜前膜形態學變化，視覺功能改變以及視網膜前膜移除手術後結果。在此研究中，我們在視網膜前組織中進行組織免疫化學染色研究以明瞭手術後視網膜前膜形成的性質。我們的研究結果顯示，玻璃體切除手術後產生的視網膜前膜是一種複雜的、含不同新生血管密度的組織。這些視網膜前膜通常的表現為廣泛分佈，進展迅速，並會引起黃斑部變形。相關的危險因素包括糖尿病視網膜病變的活動性，嚴重的纖維血管增生，玻璃體手術後出血和纖維血管的殘留。在大多數的情況下，視網膜前膜去除手術是對視力恢復是有益的，而且復發並不少見。
糖尿病病人體內的高血糖會增加了活性氧(reactive oxygen species)的產生並消耗細胞抗氧化的防禦能力，導致氧化壓力(oxidative stress)增強。慢性高氧化壓力被認為是糖尿病視網膜病變的主要原因之一。視網膜因具有高含量的不飽和脂肪酸和高氧攝取，這增加了脂質氧化和活性氧的產生。目前認為這些變化會使視網膜比任何其他組織更容易受到氧化壓力的損傷。
發炎反應可能在糖尿病視網膜病變的發展和進展中起關鍵作用。活性氧是轉錄因子核因子κB（NF-κB）的強刺激物，活性氧會增加細胞激素和趨化介素的轉錄以及增加負責一氧化氮和前列腺素E2的合成酶。所有這些因素都與糖尿病視網膜病變的進展有關。長期以來已知抗氧化劑可抑制發炎反應。在糖尿病視網膜病變的動物模型中，抗氧化劑抑制NF-κB活性，並且抑制白血球駐留和減少白血球生成誘導型一氧化氮合酶(iNOS)。此外，抗氧化劑可抑制糖尿病大鼠中無細胞微血管的形成和避免周細胞(pericytes)細胞核消失的情形。另外，抗氧化劑抑制活性氧的形成並增加抗氧化防禦酶系統的能力。因此，抗氧化劑可能會減少視網膜氧化壓力的生物損傷，減輕發炎程度，並阻止糖尿病視網膜病變的惡化。
藻紅素(astaxanthin)和葉黃素(lutein)都是含有數個雙鍵的羥基類胡蘿蔔素(hydroxycarotenoids)的葉黃素家族。他們可以有效清除活性氧，所以是強力的生物抗氧化劑和抗發炎劑。藻紅素是一種比其他類胡蘿蔔素更有效的抗氧化劑，包括葉黃素，β-胡蘿蔔素，角黃素和玉米黃素。在我們的研究中，我們證明了葉黃素及藻紅素可能經由調節NF-κB活性，減少了糖尿病大鼠模型中的眼部氧化壓力和發炎現象，而有神經保護作用。
西洛他唑(cilostazol)是一種磷酸二酯酶3型(phosphodiesterase 3)抑制劑， 它可以增加細胞內環腺苷一磷酸的濃度，進而減少血小板內的鈣離子，導致抑制血小板聚集及增進血管擴張，最後降低動脈內血壓。有人進一步研究發現西洛他唑(cilostazol)可以減少發炎，改善微循環，而達到對視網膜神經細胞的保護作用。其機轉可能是抑制阿爾法型腫瘤壞死因子(tumor necrosis factor alpha; TNFα)造成的NF-κB活化及其下游的發炎基因的表現。
在我們的研究系列中，我們最初根據增殖性糖尿病視網膜病變的發病機制，發展了新的手術技術來治療糖尿病玻璃體出血。 隨後，我們經由組織切片研究了增殖性糖尿病視網膜病變的獨特組織病理學特徵。最新動物實驗研究顯示，藻紅素及西洛他唑(cilostazol)都是高效抗氧化劑，可以用於預防糖尿病視網膜病變進展並保護眼部功能。

關鍵字

視網膜前部冷凍治療 ； 藻紅素 ； 癌思停 ； 西洛他唑 ； 血管纖維化增生 ； 氧化壓力 ； 活性氧

並列摘要

Pars plana vitrectomy (PPV) has been one of the major treatments for various complications of proliferative diabetic retinopathy (PDR). With modern instruments and techniques, the anatomical success rate has been quite high, ranging from 70% to 90%, depending upon the severity of the disease process. Post-operatively, about one third of successfully treated patients develop recurrent vitreous hemorrhage (VH). The bleeding may come from residual fibrovascular tissues or may be related to peripherally located postoperative neovascularization. Recurrent VH not only slows visual recovery but also jeopardizes an otherwise successful operation. Therefore, it is crucial to execution of proper preventive managements for reducing intra- and postoperative preretinal hemorrhage in complicated diabetic vitrectomy (DV). If recurrent vitreous hemorrhage occurs postoperatively, appropriate managements might induce regression of neovasculariation and reduce the possibility of repeated bleeding, thus shortening the reabsorption time and reducing the need for surgery. In our serial studies, we designed several clinical studies and developed some effective surgical methods including (1)intravitreal bevacizumab (1.25 mg) injection 1 week before surgery combined primary silicone oil infusion; (2) combination treatment of anterior retinal cryotherapy (ARC) & intravitreal bevacizumab injection (1.25 mg) and (3) repeated intravitreal bevacizumab injection for post-diabetic vitrectomy VH patients to prevent post-diabetic vitrectomy hemorrhage and to treat post-vitrectomy diabetic vitreous hemorrhage. Our results revealed that these methods were effective and safe for treating recurrent diabetic VH.
In PDR, diabetic fibrovascular proliferation (FVP) is unique for its complex vitreoretinal relationship, the frequent presence of epiretinal membrane (ERM) and the strong proinflammatory and pro-angiogenic environment might resulted in PDR.
ERM formation may occur in various clinical settings. It results from cellular proliferation along the surface of the retina, but the origin of these proliferating cells has been debated. Intraretinal glial cells, other retinal connective tissue cells, and even retinal pigment epithelial cells have been found in surgical specimens of ERM. The pathophysiology of ERM formation may vary widely in different disease conditions. In diabetic retinopathy, ERM may occur before surgery, or develop after successful vitrectomy. Specific features in the post-vitrectomized eye, such as ischemia, inflammation, hemorrhage, and residual fibrovascular membrane, may predispose the eye to ERM formation. To better understand the effect of the ERM on the macular structure and prognosis after its removal in eyes that have undergone DV, a retrospective study was conducted to evaluate the possible risk factors, morphological changes, visual function changes, and the surgical results of ERM after DV. In addition, immunohistochemical studies were performed in preretinal tissues to elucidate the nature of ERM formation after DV. Our results showed the post-DV ERM is a complex tissue with variable vascularity that often presents with widespread distribution, rapid progression, and causes macular distortion. Associated risk factors include active PDR, high FVP grade, post-DV hemorrhage, and residual fibrovascular stumps. Membrane removal surgery may be beneficial in selected cases, but recurrence is not uncommon.
The hyperglycemia that occurs in diabetes increases the production of reactive oxygen species (ROS) and depletes cellular antioxidant defense capacities, resulting in enhanced oxidative stress. Chronic oxidative stress is considered one of the primary causes of diabetic retinopathy. The retina has a high content of unsaturated fatty acids and high oxygen uptake, which increases lipid oxidation and ROS production. This is commonly thought to make the retina more vulnerable than any other tissue to oxidative stress damage.
Inflammation may also play a key role in the development and progression of diabetic retinopathy. ROS are strong stimulators of the transcription factor nuclear factor kappa B (NF-κB), which increases the transcription of inflammatory cytokines and chemokines as well as enzymes responsible for nitric oxide and prostaglandin E2 synthesis. All of these factors are involved in the pathogenesis of diabetic retinopathy. Antioxidants have long been known to inhibit inflammatory responses. In animal models of diabetic retinopathy, antioxidants inhibit NF-κB activity, and reduce leukostasis and leukocyte expression of inducible nitric oxide synthase. Moreover, antioxidants can inhibit the formation of cell-free capillaries and generation of pericyte ghosts in diabetic rats. In addition, antioxidants inhibit the formation of ROS and increase the capabilities of the antioxidant defense enzyme system. Therefore, antioxidants might diminish the biologic damage of oxidative stress in the retina, abate the level of inflammation, and arrest the progression of diabetic retinopathy.
Astaxanthin (AST) and lutein both are from the xanthophyll family of hydroxycarotenoids which contain several double bonds. They could scavenge ROS to be powerful biological antioxidants and anti-inflammatory agents. AST is a more potent antioxidant than other carotenoids, including lutein, β-carotene, canthaxanthin, and zeaxanthin. In our study, we proved the xanthophyll carotenoid AST had neuroprotective effects and would reduce ocular oxidative stress, and inflammation in the STZ diabetic rat model, which might be mediated by downregulation of NF-κB activity.
Cilostazol is a phosphodiesterase 3 (PDE3) inhibitor which has been shown to increase intracellular cyclic adenosine monophosphate levels and then decrease intracellular Ca2+ levels in platelets, resulting in inhibition of platelet aggregation and vasodilation leading to a reduction in arterial pressure. It has been shown to reduce inflammation and improve microcirculation to achieve neuroprotective effects in the retina by inhibiting tumor necrosis factor alpha (TNFα)-induced NF-κB activation and proinflammatory gene expressions.
In our study series, we initially developed new surgical techniques according to the pathogenesis of PDR to treat diabetic vitreous hemorrhage. Subsequently, we studied the unique histopathological features of PDR by histologic sections. In the latest study, we applied AST and cilostazol, both are potent antioxidants, to prevent diabetic retinopathy progressing and to preserve ocular functions.

並列關鍵字

anterior retinal cryotherapy ； astaxanthin ； bevacizumab ； cilostazol ； diabetic retinopathy ； fibrovascular proliferation ； oxidative stress ； reactive oxygen species

參考文獻

1. Giacco F, Brownlee M. Oxidative stress and diabetic complications. Circ Res 2010;107:1058-1070.

2. Baynes JW, Thorpe SR. Role of oxidative stress in diabetic complications: a new perspective on an old paradigm. Diabetes 1999;48:1-9.

3. Kaneto H, Katakami N, Kawamori D, et al. Involvement of oxidative stress in the pathogenesis of diabetes. Antioxid Redox Signal 2007;9:355-366.

4. Pan HZ, Zhang H, Chang D, et al. The change of oxidative stress products in diabetes mellitus and diabetic retinopathy. Br J Ophthalmol 2008;92:548-551.

5. Giugliano D, Ceriello A, Paolisso G. Oxidative stress and diabetic vascular complications. Diabetes Care 1996;19:257-267.