scopolamine-induced tissue damage assessment in dry eye rat models

ラットにおけるスコポラミン誘発性涙腺機能障害

By 2021, approximately 12% of people are significantly affected by dry eyes1, making it one of the most common chronic eye diseases. Dry eye can be divided into two types: aqueous-deficient dry eye (ADDE) and evaporative dry eye (EDE)2, depending on the different factors that affect the disease. ADDE is further divided into Sj&#246;gren&#39;s syndrome (SS) and non-SS, but the majority of dry eye patients are non-SS patients in clinical3. Chronic dry eye symptoms seriously affect the visual quality of patients. Currently, the conventional treatment of DED involves the application of artificial tears to lubricate the ocular surface and physical therapy of the eyelids. However, dry eye syndrome may not offer a complete cure in many cases. Therefore, studying the pathogenesis of dry eye disease is crucial for the development of new therapies and drugs. Animal models of dry eye syndrome provide a foundation for further research.
There are many ways to construct animal models of dry eye syndrome4, including changing tear secretion levels by altering hormone levels. For example, removing the testes of rats can reduce androgen secretion, increase tear secretion, and decrease the concentration of free secretory component (SC) and IgA in tears5,6. Another method is to indicate autoimmune reactions in the lacrimal gland by removing the eye surface nerves that control the gland. Additionally, directly reducing tear secretion can be achieved by surgically removing the lacrimal gland7. Changing environmental conditions can also accelerate tear evaporation. For example, culturing animals in low humidity and dry ventilation conditions can establish a model of excessive evaporative dry eye8, which can be combined with other methods to increase the severity of dry eye. The main drugs used to induce dry eye experimental models are atropine and scopolamine9. As parasympathetic inhibitors, both can induce pharmacological blockade of cholinergic (muscarinic) receptors in the lacrimal gland and inhibit tear secretion. Compared with dry eyes caused by atropine muscle injection10, scopolamine has a stronger inhibitory effect on secretion glands, a longer duration of drug action, and weaker effects on cardiac, small intestinal, and bronchial smooth muscles. It is one of the most mature drugs for dry-eye animal models.
Different methods can be used to induce dry eye with scopolamine, such as subcutaneous injection, drug pump, or patch application4,11,12. In order to reduce the frequency of drug administration to experimental animals, many researchers apply transdermal patches to the tails of mice or use drug pumps. However, both of these methods have limitations. For example, the absorption of transdermal patches needs to take into account the individual absorption of mice, which can lead to inconsistent drug dosage. Although drug pumps can accurately control the dosage of each administration, they are not always compatible with the drug being delivered or the concentration being used. They also need to be placed surgically &#8211; which is more invasive to the animal, requiring an anesthetic event, and there is potential for post-surgical complications such as dehiscence. Subcutaneous injection, although more cumbersome, can ensure accurate dosage for each administration and maintain consistency in drug administration among different rats. At the same time, it has a lower cost and is suitable for conducting a large number of animal experiments.
This study applies repeated subcutaneous injection of scopolamine to establish a dry eye rat model. We analyze dry eye indicators such as corneal defects, tear secretion levels, and pathological morphology of the cornea, conjunctiva, and lacrimal gland. By combining drug concentration, pathological manifestations, and dry eye symptoms, we further elaborate on the dry eye rat model in detail, providing more accurate experimental data for the study of dry eye treatment and pathological mechanisms. We also describe the modeling process in detail for future researchers.

著者スポットライト:ドライアイ動物モデルの開発における課題と今後の研究の方向性

スコポラミンによる涙腺機能障害を有するラットドライアイモデル

The process of developing a dry eye animal model is a long one. It requires continuous subcutaneous injection of scopolamine in the hind legs of rice to suppress the secretion of the lacrymal gland in rice. So, experimenters need to have enough patience to administer the medication regularly over a long period of time.Our protocol is a very useful method that can created different degrees of dry eye models based on different drug concentrations. It can make it easiest for the future researchers to study the treatment of dry eye disease. Our team will continue to explore the field of lacrimal gland regeneration, very excited to work on reconstructing 3D and conducting transplantation experiments.Our ultimate goal is to develop more advanced and efficient treatment options for individuals suffering from dry eye syndrome.

ドライアイラットモデルにおけるスコポラミン誘発性組織損傷評価

まず、ネズミの体をしっかりと持ち、後ろ足を伸ばします。注射部位をアルコールで拭きます。次に、親指と指の間の皮膚のひだの付け根に26ゲージの針を備えた1ミリリットルの注射器を挿入します。シリンジプランジャーを引いて針を吸引します。必要に応じて針を取り外して位置を変えます。0.5ミリリットルの0.9%塩化ナトリウムをスコポラミン臭化水素酸塩の有無にかかわらず、1回の定常運動で注射します。.涙液分泌試験を行うには、まず濾紙ストリップの半分を中心線に沿ってカットします。ストリップの頭をトリミングして滑らかにします。ラットの下まぶたの結膜嚢の外側3分の1にろ紙ストリップを置きます。5分後、ピンセットを使用してろ紙ストリップをマイクロ遠心チューブにクランプします。チューブの壁にマークを付けて、涙の量を記録します。ラットの目の角膜を染色するには、0.5マイクロリットルの0.5%フルオレセインナトリウム溶液を各眼の下結膜嚢に滴下します。3分後、青色光の下で角膜を観察します。安楽死させたネズミの目から球結膜を取ります。各ラットの右側から角膜を切除し、すぐに4%パラホルムアルデヒドで固定します。耳と目尻を結ぶ線に沿って皮下組織から頭側表皮を切り取ります。次に、切開を両側に広げ、黄色がかった余分な眼窩腺を分離します。次に、ラットの毛皮を完全に取り除き、0.9%塩化ナトリウム溶液で余分な眼窩腺を分離します。スコポラミン群の涙液分泌は有意に減少した。スコポラミンは、角膜染色の欠如によって証明されるように、角膜上皮欠損を引き起こさなかった。スコポラミン群の角膜上皮は対照群よりも比較的薄かった。また、7.5群では、基底層に緩やかな細胞内接続と空胞構造が見られました。対照群は角膜上皮が有意に厚かった。スコポラミンは涙腺に炎症性変化を引き起こし、機能的な涙腺の損傷を示しているようでした。.グループ2.5とグループ5では粘液性物質の減少が観察され、遊離リンパ球浸潤が見られました。さらに、グループ7.5では、不規則な形状の上皮細胞を持つ細胞の不規則な配置が示されました。スコポラミンはまた、結膜の形態学的変化を引き起こしました。.

Research

JoVE Journal

Medicine

Scopolamine-Induced Tissue Damage Assessment in Dry Eye Rat Models

A Rat Dry Eye Model with Lacrimal Gland Dysfunction Induced by Scopolamine

Aqueous-deficient dry eye (ADDE) is a type of dry eye disease that can result in the reduction of tear secretion quantity and quality. Prolonged abnormal ...

医学