We thank Dr. John Chang (Radiation Oncology, LSU Health Science Center) for assistance with radiation dose measurements. The study was supported by the American Cancer Society (Grant number: 116945-RSG-09-038-01-CCE), National Institute of Health (Grant number: R21CA173162) and the Feist-Weiller Cancer Center.

すべての手順はLSU健康、シュリーブポート、動物実験委員会によって承認され、実験動物の管理と使用のためのNIHガイドラインに従ってありました。 ラット顎下唾液腺の1カニューレ挿入<ol><li> シリンジ-チューブアセンブリの調製 <ol><li>メスでPE10ポリエチレンチューブの10cmの長さをカットします。人差し指と親指の間のチューブの両端を持ちます。穏やかな炎上記チューブの中央部分を熱し、ゆっくり両側を引っ張ることにより、その長さを倍増させる軟化チューブを伸ばします。 </li><li>先細端部と各- 2カニューレを取得するために45 Oの角度でメスで途中でチューブをカットします。 0.5ccのインスリン注射器の29 Gの針の上にフィットさせることによって、カニューレの非テーパ状の端部を展開します。 </li><li>カニューレを外し、シリンジに生理食塩水の220μLを描きます。気泡を取り除くために、タップします。 cannulを取り付け針の上に、そして空気を排出し、カニューレを介して、溶液の自由な流れを確保するために、シリンジのプランジャーを押してください。 200μlに音量を調整します。 </li></ol></li><li> 顎下腺ダクトの口腔内カニューレ挿入 注：オートクレーブ楽器は手続きの前に、ホットビーズ滅菌器の手順の間に、滅菌します。 <ol><li>スプラーグドーリーラットを計量し、皮下注射器で計算されたケタミンの量（42 mgの/キログラム）/キシラジン（8mg / kg）/アセプロマジン（1.4ミリグラム/キログラム）のミックスを分注します。 </li><li>人差し指の間尾と片手の親指の付け根を把握し、それを把握するために身体の上にもう一方の手をスライドさせることにより、動物を拘束。親指と残りの指で胴体を保持しながら、頭部の側面に沿って人差し指と中指を立てます。 </li><li>後肢の筋肉組織に麻酔薬を注入します。つま先のピンチと眼瞼反射によって麻酔の深さを確認してください。乾きしばらくのを防ぐために目の潤滑剤を塗布動物は麻酔下です。 </li><li>特別に設計されたプラットフォーム（ 図1）に動物を置き、横バーの上側切歯に係合します。下の切歯の周りにゴムバンドをループし、プラットフォームにそれを固定することにより、下顎をプルダウン。 </li><li>舌を通して滅菌縫合糸を渡し、口の床を上げるためにそれを持ち上げます。止血鉗子で縫合糸をクランプ、および横方向のバーの上にそれを渡します。 </li><li>カスタムビルドの頬スプレッダー（ 図1）と頬を展開し、解剖顕微鏡下で、口の床の上に舌下乳頭を探します。 </li><li>繊細な鉗子を用いて行うPE10チューブのテーパ状の端部をつかみます。そっと舌下乳頭上の腺管開口部にカニューレの先端を操作します。それは3ねじ込むことによって、カニューレの配置を確認してください - ダクトに5ミリメートルを。それは、任意の障害物なしに通過することを確認してください。 </li></ol></li><li> 顎下腺Instillation <ol><li>首筋に皮下アトロピン（0.5ミリグラム/キログラム）を注入し、唾液分泌の減少のために10分を待ちます。 </li><li>シアノアクリレート（接着剤）の低下にダクト開口部にカニューレを固定し、乾燥させます。ゆっくり〜50マイクロリットル/分の速度でシリンジプランジャを押して腺（200μL/腺）溶液を植え付けます。 </li><li>止血鉗子でチューブをつぶす、と慎重に注射器を取り外します。動物が意識を取り戻すまで60分 - 30用のダクト内チューブを保持します。舌を保持している縫合糸を外します。 </li><li>別々のケージに動物を移し、リカバリ中にそれを暖かく保つために加熱ランプを使用しています。それは胸骨横臥を維持するために意識を取り戻したまで無人の動物を放置しないでください。 </li><li>動物が完全に歩行した後、食物および水に無制限にアクセスできるビバリウムでそれを収容します。 </li></ol></li></ol> 2.ローカライズ分割照射顎下腺の<ol><li>前述したように動物を拘束し、後肢でケタミン（33 mgの/キログラム）/キシラジン（6ミリグラム/キログラム）/アセプロマジン（1ミリグラム/キログラム）ミックスの筋肉内投与で麻酔。麻酔の深さを確認し、目に潤滑剤を塗布。 </li><li>線形加速器卓上に動物仰向けに置き、頭を傾けることによって、その首を拡張します。胸骨の上部に下顎骨の下縁からの領域を包含するように照射野（3センチスリット幅）をコリメートします。 </li><li>領域の上に1cmの組織等価ボーラスを配置し、放射線源100センチボーラスの頂部との間の距離を調整します。 </li><li>線形加速器の6 MV光子ビームを用いて動物（2.5 Gyの）を照射します。線量率、視野サイズ、及び放射線源からのボーラスの表面までの距離は、露光時間を決定するであろう。現在のセットアップでは、動物は約1グレイ/分の線量率で照射しました。 </li><li>露出を繰り返します。 2.5 Gyの/ 8日目の合計日々;間で2日間隔で4日/週。リカバリ中に動物を暖かく保ちます。それは完全に歩行した後に飼育箱に動物を転送します。 </li><li>腺機能14を測定するために、8週間照射後刺激顎下腺の唾液を収集します。冷たい4％パラホルムアルデヒド/リン酸緩衝生理食塩水のpHが7.2の心臓灌流によって麻酔下で動物を安楽死させます。組織学的および免疫組織化学14を分析するための顎下腺を摘出。 </li></ol>

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The authors have nothing to disclose.

唾液腺は、多くの場合、頭頸部癌、首ノードの選択科目切除、または地域の血液悪性腫瘍に対する放射線治療を受けた患者における組織回復のしきい値を超えた放射線量を受けます。腺の流体を分泌する腺房細胞が最終的に分化しているが、それらは逆説的に放射線に敏感です。分泌機能は、放射線の最初の数週間以内に低下し、慢性低唾液出力の不可逆的な腺の損傷をもたらします。貧し?...

健康な組織の担保破壊は、癌治療の有害な副作用の数を占めています。放射線分野でうそをつく大唾液腺の一部または全部は、必然的に破壊されます。従って、骨髄移植の前頭部および頸部癌、子宮頸癌、リンパ腫、または全身照射のための放射線治療を受けている多くの患者は、放射線、唾液腺機能低下2-6の最も一般的かつ永続的な副作用の一つを被ります。 唾液腺の流体生産腺房細胞は、放射線への急性敏感です。唾液腺の損傷は、唾液機能低下と呼ばれる状態を唾液の流れの大幅な逓減を引き起こします。唾液の流れにおける慢性の減少は、咀嚼、嚥下、スピーチ、そして味が、激しい痛み、粘膜涙、嚥下障害、日和見感染症の病的後遺症などの重要経口活動を阻害し、虫歯が悪化します患者の幸福と機能2,3。 放射線治療関連の唾液細胞の損失は不可逆的であるため、口腔乾燥症のない矯正治療はありません。人工唾液代用とprosecretory薬で症状を和らげに焦点を当て現在の治療は長期の救済6には無効です。改良された放射線送達技術は、症状の重症度を減少させる助けているが、正常組織毒性とその合併症は、がん治療6,7の制限要因のまま。放射線治療に関連する合併症を予防するための先制措置は、従って、標準になりつつあります。フリーラジカル酸素種を除去ラジオ保護剤、里親細胞再増殖、またはDNA修復を高めるためには、唾液機能低下8-11を回避するために探索されています。 外分泌唾液腺の分泌物は、主排泄ダクトを介して口の中に排出します。目の口腔内挿管造影剤の注入のための電子排泄ダクトは外来処置として日常的に行われます。同様のアプローチを利用して、唾液腺に直接局所治療12の対象とすることができます。別に全身副作用のリスクを減少させるから、retroductal腺点滴は利益を追加しました。乳管ツリーの周りの唾液細胞の単層の配置は、すべての唾液腺上皮細胞をターゲットにすることを可能にし、不要な治療拡散を低減するための障壁として腺行為の繊維状のカプセル化。本質的には、唾液腺は、このような放射線誘発唾液機能低下などの腺苦痛の標的治療のために最適に適しています。 週間の期間のための - （2.5 Gyの/分数/日、5日の週1.8）癌治療のための従来の放射線が少量で提供されています。したがって、実験モデルにおいて長引く放射スキームに対する有効性を示している放射線防護の治療は、より大きな臨床ベアリングを持っています。 COMPRO分別放射線は小動物に記録されたが、放射線源、用量分画、および使用されるプロトコルは9,10,13変化された後、唾液機能mised。 このレポートでは、患者関連の放射線源と線量分画を用いてラット顎下腺の放射線へのローカライズさretroductal送達のための方法を確立します。

<table><tbody><tr><td>Intramedic Polyethylene tubing (PE10)</td><td>Becton Dickson</td><td>427401</td><td></td></tr><tr><td>1/2 cc Insulin Syringe U-100</td><td>Becton Dickson</td><td>309306</td><td></td></tr><tr><td>Artificial Tears</td><td>Miller Vet Supply&amp;nbsp;</td><td>5098-9840-64</td><td></td></tr><tr><td>Hot Bead Sterilizer</td><td>Fine Science Tools</td><td>18000-45</td><td></td></tr><tr><td>Perma-Hand silk suture</td><td>Ethicon</td><td>K833H</td><td></td></tr><tr><td>Graefe forcep</td><td>Fine Science Tools</td><td>11051-10</td><td></td></tr><tr><td>Olympus SZX16 Stereo Microscope</td><td>Hunt Optics and Imaging</td><td></td><td></td></tr><tr><td>6 MV Linear Accelerator</td><td>Elekta</td><td></td><td></td></tr><tr><td>Bolus - Skinless&amp;nbsp;</td><td>Civco</td><td>MTCB410</td><td></td></tr><tr><td>Heat Lamp</td><td>Braintree Scientific</td><td>HL-1&amp;nbsp;110V</td><td></td></tr></tbody></table>

retroductal submandibular gland instillation and localized fractionated irradiation in a rat model of salivary hypofunction

Normal tissues that lie within the portals of radiation are inadvertently damaged. Salivary glands are often injured during head and neck radiotherapy. Irreparable cell damage results in a chronic loss of salivary function that impairs basic oral activities, and increases the risk of oral infections and dental caries. Salivary hypofunction and its complications gravely impact a patient's comfort. Current symptomatic management of the condition is ineffective, and newer therapies to assuage the condition are needed. 
Salivary glands are exocrine glands, which expel their secretions into the mouth via excretory ducts. Cannulation of these ducts provides direct access to the glands. Retroductal delivery of a contrast agent to major salivary glands is a routine out-patient procedure for diagnostic imaging. Using a similar procedure, localized treatment of the glands is feasible. However, performing this technique in preclinical studies with small animals poses unique challenges. In this study we describe the technique of retroductal administration in rat submandibular glands, a procedure that was refined in Dr. Bruce Baum's laboratory (NIH)1, and lay out a procedure for local gland irradiation.

低侵襲性唾液腺撮影技術、大唾液腺の局所治療を適応させることが可能です。ラット顎下唾液腺におけるRetroductal投与はウォートンのダクトの口腔内挿管（ 図2）により実行しようとしました。ウォートンの唾液管は、口の階にある舌乳頭に開くが、オリフィスは容易には見えません。カニューレの挿入は、従って、穏やかなプロービングすることによっ?...

Watch this Scientific Journal Video about Retroductal Submandibular Gland Instillation and Localized Fractionated Irradiation in a Rat Model of Salivary Hypofunction at JoVE.com

Retroductal Submandibular Gland Instillation and Localized Fractionated Irradiation in a Rat Model of Salivary Hypofunction

Salivary gland hypofunction, a major adverse effect of head and neck radiotherapy diminishes a patient's quality of life. The demonstration of efficacy of new therapies in animal models is a prerequisite before clinical transition. This protocol describes retroductal administration and local irradiation of rat submandibular glands.

唾液機能低下のラットモデルにおけるRetroductal顎下腺点滴とローカライズされた分割照射

Normal tissues that lie within the portals of radiation are inadvertently damaged. Salivary glands are often injured during head and neck radiotherapy. ...

retroductal-submandibular-gland-instillation-localized-fractionated

Universidad de Cartagena UDEC

Research

JoVE Journal

Medicine

11.3K ビュー.  Louisiana State University Health Sciences Center. Salivary gland hypofunction, a major adverse effect of head and neck radiotherapy diminishes a patient's quality of life. The demonstration of efficacy of new therapies in animal models is a prerequisite before clinical transition. This protocol describes retroductal administration and local irradiation of rat submandibular glands.

ビデオ: Retroductal Submandibular Gland Instillation and Localized Fractionated Irradiation in a Rat Model of Salivary Hypofunction

Cannulation of the Mouse Submandibular Salivary Gland via the Wharton's Duct

Severe salivary gland hypofunction is frequently found in patients with Sj&ouml;gren's syndrome and those who receiving therapeutic 
irradiation in their head and neck regions for cancer treatment. Both groups of patients experience symptoms such as xerostomia (dry mouth), dysphagia 
(impaired chewing and swallowing), severe dental caries, altered taste, oro-pharyngeal infections (candidiasis), mucositis, pain and discomfort.
One innovative approach of regenerative medicine for the treatment of salivary gland hypo-function is speculated in RS Redman, E Mezey 
et al. 2009: stem cells can be directly deposited by cannulation into the gland as a potent method in reviving the functions of the impaired organ. Presumably, 
the migrated foreign stem cells will differentiate into glandular cells to function as part of the host salivary gland. Also, this cannulation technique is an 
expedient and effective delivery method for clinical gene transfer application.
Here we illustrate the steps involved in performing the cannulation procedure on the mouse submandibular salivary gland via the Wharton's duct 
(Fig 1). C3H mice (Charles River, Montreal, QC, Canada) are used for this experiment, which have been kept under clean conventional conditions at 
the McGill University animal resource center. All experiments have been approved by the University Animal Care Committee and were in accordance with the 
guidelines of the Canadian Council on Animal Care.
For this experiment, a trypan blue solution is infused into the gland through the opening of the Wharton's duct using a 
insulin syringe with a 29-gauge needle encased inside a polyethylene tube. Subsequently, the mouse is dissected to show that the infusions migrated 
into the gland successfully.

A protocol for the cannulation of the mouse submandibular salivary gland via the Wharton's duct is described. For this experiment, the trypan blue solution is used as a dyer to demonstrate how this technique effectively delivers infusions into the targeted gland, and to suggest the reliability of this new approach as a potential clinical drug/cell therapy for the regeneration of salivary glands.

ワルトン管を経由してマウス顎下唾液腺のカニュレーション

Severe salivary gland hypofunction is frequently found in patients with Sj&ouml;gren's syndrome and those who receiving therapeutic 
irradiation in their ...

医学

Isolation of Mouse Salivary Gland Stem Cells

Mature salivary glands of both human and mouse origin comprise a minimum of five cell types, each of which facilitates the production and excretion of saliva into the oral cavity. Serous and mucous acinar cells are the protein and mucous producing factories of the gland respectively, and represent the origin of saliva production. Once synthesised, the various enzymatic and other proteinaceous components of saliva are secreted through a series of ductal cells bearing epithelial-type morphology, until the eventual expulsion of the saliva through one major duct into the cavity of the mouth. The composition of saliva is also modified by the ductal cells during this process.
In the manifestation of diseases such as Sj&ouml;gren's syndrome, and in some clinical situations such as radiotherapy treatment for head and neck cancers, saliva production by the glands is dramatically reduced 1,2. The resulting xerostomia, a subjective feeling of dry mouth, affects not only the ability of the patient to swallow and speak, but also encourages the development of dental caries and can be socially debilitating for the sufferer.
The restoration of saliva production in the above-mentioned clinical conditions therefore represents an unmet clinical need, and as such several studies have demonstrated the regenerative capacity of the salivary glands 3-5. Further to the isolation of stem cell-like populations of cells from various tissues within the mouse and human bodies 6-8, we have shown using the described method that stem cells isolated from mouse salivary glands can be used to rescue saliva production in irradiated salivary glands 9,10. This discovery paves the way for the development of stem cell-based therapies for the treatment of xerostomic conditions in humans, and also for the exploration of the salivary gland as a microenvironment containing cells with multipotent self-renewing capabilities.

An optimized protocol for the isolation of stem cells from the mouse salivary gland is described. The method employs enzymatic and mechanical digestion, and permits isolation of salispheres containing cells with characteristics of stem cells.

マウス唾液腺幹細胞の単離

Mature salivary glands of both human and mouse origin comprise a minimum of five cell types, each of which facilitates the production and excretion of ...

生物学

Retroductal Nanoparticle Injection to the Murine Submandibular Gland

Two common goals of salivary gland therapeutics are prevention and cure of tissue dysfunction following either autoimmune or radiation injury. By locally delivering bioactive compounds to the salivary glands, greater tissue concentrations can be safely achieved versus systemic administration. Furthermore, off target tissue effects from extra-glandular accumulation of material can be dramatically reduced. In this regard, retroductal injection is a widely used method for investigating both salivary gland biology and pathophysiology. Retroductal administration of growth factors, primary cells, adenoviral vectors, and small molecule drugs has been shown to support gland function in the setting of injury. We have previously shown the efficacy of a retroductally injected nanoparticle-siRNA strategy to maintain gland function following irradiation. Here, a highly effective and reproducible method to administer nanomaterials to the murine submandibular gland through Wharton's duct is detailed (Figure 1). We describe accessing the oral cavity and outline the steps necessary to cannulate Wharton's duct, with further observations serving as quality checks throughout the procedure.

Local drug delivery to the submandibular glands is of interest in understanding salivary gland biology and for the development of novel therapeutics. We present an updated and detailed retroductal injection protocol, designed to improve delivery accuracy and experimental reproducibility. The application presented herein is the delivery of polymeric nanoparticles.

マウス顎下腺への Retroductal ナノ粒子注入

Two common goals of salivary gland therapeutics are prevention and cure of tissue dysfunction following either autoimmune or radiation injury. By locally ...

生物工学

A Method for the Measurement of Salivary Gland Function in Mice

Patients with Sj&#246;gren&#39;s syndrome, an autoimmune disease affecting the exocrine glands, develop salivary gland inflammation and have reduced saliva production. Similarly, saliva production is severely compromised in patients receiving radiation treatment for head and neck cancers. Rodent models, developed to mimic these clinical conditions, facilitate an understanding of the disease pathogenesis and allow for the development of new therapeutic strategies. Therefore, the ability to accurately, reproducibly, and repeatedly measure salivary gland function in animal models is critical. Building on procedures previously described in the literature, a method was developed that meets these criteria and was used to evaluate salivary gland function in mice. An additional advantage of this new method is that it is easily mastered, and has little inter-operator variation. Salivary gland function is evaluated as the amount (weight or volume) or rate (mL/min) of saliva produced in response to pilocarpine stimulation. The collected saliva is a good source for the analyses of protein content, immunoglobulin concentrations, and other biomolecules.

Salivary gland hypofunction is a frequent consequence of autoimmune disease and radiation therapy. Reproducible evaluation of salivary gland function in mouse models of these diseases is a technical challenge. Here, a simple method for accurate and reproducible measurement of saliva production in mice is described.

マウスの唾液腺機能の測定法

Patients with Sj&#246;gren&#39;s syndrome, an autoimmune disease affecting the exocrine glands, develop salivary gland inflammation and have reduced ...

免疫・感染学

Preparation of Murine Submandibular Salivary Gland for Upright Intravital Microscopy

The submandibular salivary gland (SMG) is one of the three major salivary glands, and is of interest for many different fields of biological research, including cell biology, oncology, dentistry, and immunology. The SMG is an exocrine gland comprised of secretory epithelial cells, myofibroblasts, endothelial cells, nerves, and extracellular matrix. Dynamic cellular processes in the rat and mouse SMG have previously been imaged, mostly using inverted multi-photon microscope systems. Here, we describe a straightforward protocol for the surgical preparation and stabilization of the murine SMG in anesthetized mice for in vivo imaging with upright multi-photon microscope systems. We present representative intravital image sets of endogenous and adoptively transferred fluorescent cells, including the labeling of blood vessels or salivary ducts and second harmonic generation to visualize fibrillar collagen. In sum, our protocol allows for surgical preparation of mouse salivary glands in upright microscopy systems, which are commonly used for intravital imaging in the field of immunology.

We describe a protocol to surgically expose and stabilize the murine submandibular salivary gland for intravital imaging using upright intravital microscopy. This protocol is easily adaptable to other exocrine glands of the head and neck region of mice and other small rodents.

マウス顎下腺唾液腺の直立した生体顕微鏡検査のための準備

The submandibular salivary gland (SMG) is one of the three major salivary glands, and is of interest for many different fields of biological research, ...

Ultrasound-guided Botulinum Toxin-A Injections: A Method of Treating Sialorrhea

Neurological diseases can be complicated by sialorrhea, an excessive flow of saliva. Patients suffering from moderate to severe sialorrhea have an impaired quality of life, often worsened by correlated complications such as aspiration pneumonia, oral infections, dental caries, and maceration of the skin. Diverse therapeutic approaches have been proposed for the treatment of sialorrhea, including surgery and the use of anticholinergic agents, with limited results and the possible occurrence of serious adverse events. Recently, botulinum toxin (BoNT) injection within the major salivary glands has been proposed in patients refractory to anticholinergic therapy, with the aim of inhibiting local acetylcholine release and gland activity. 
In order to obtain a better outcome in terms of reduction of saliva production, efficacy, duration, and avoidance of major adverse events, we developed an ultrasound-guided BoNT-type A injection technique accurately described in the text. Here we present a method of treating sialorrhea with bilateral parotid and submandibular gland BoNT-type A injections under ultrasound guidance. Four quadrants of the parotid gland and two quadrants of the submandibular gland are visualized and injected using two accesses and one access, respectively. 
The ultrasound-guided procedure provides a simple, non-invasive, real-time visualization of the muscular and glandular tissues and their surrounding structures, optimizing treatment efficacy and safety.

Botulinum toxin is injected into the salivary glands for the treatment of sialorrhea. The ultrasound-guided botulinum toxin injection in both the parotid and submandibular glands demonstrates a sustained efficacy and the absence of serious side effects.

ボツリヌス毒素Aの注射を超音波誘導：唾液分泌過多を治療する方法

Neurological diseases can be complicated by sialorrhea, an excessive flow of saliva. Patients suffering from moderate to severe sialorrhea have an ...

Murine Salivary Functional Assessment via Pilocarpine Stimulation Following Fractionated Radiation

Hyposalivation is commonly observed in the autoimmune reaction of Sj&#246;gren's syndrome or following radiation injury to the major salivary glands. In these cases, questions remain regarding disease pathogenesis and effective interventions. An optimized technique that allows functional assessment of the salivary glands is invaluable for investigating exocrine gland biology, dysfunction, and therapeutics. Here, we present a step by step approach to performing pilocarpine stimulated saliva secretion, including tracheostomy and the dissection of the three major murine salivary glands. We also detail the appropriate murine head and neck anatomy accessed during these techniques. This approach is scalable, allowing for multiple mice to be processed simultaneously, thus improving the efficiency of the work flow. We aim to improve the reproducibility of these methods, each of which has further applications within the field. In addition to saliva collection, we discuss metrics for quantifying and normalizing functional capacity of these tissues. Representative data are included from submandibular glands with depressed salivary gland function 2 weeks following fractionated radiation (4 doses of 6.85 Gy).

We present a detailed approach to performing saliva collection, including murine tracheostomy and the isolation of three major salivary glands.

ピロカルピン刺激次を介してマウス唾液機能的アセスメント分別放射線

Hyposalivation is commonly observed in the autoimmune reaction of Sj&#246;gren's syndrome or following radiation injury to the major salivary glands. In ...

Retrograde Parotid Gland Infusion through Stensen's Duct in a Non-Human Primate for Vectored Gene Delivery

Salivary glands are an attractive tissue target for gene therapy with promising results already leading to human trials. They are inherently capable of secreting proteins into the bloodstream and are easily accessible, making them potentially superior tissue sites for replacement hormone production or vaccination by gene transfer. Suggested methods for gene delivery include transcutaneous injection and retrograde infusion through salivary ducts. We demonstrate how to perform Retrograde Salivary Gland Infusion (RSGI) in non-human primates. We describe the important anatomic landmarks including identification of the parotid papilla, an atraumatic method of cannulating and sealing Stensen&#39;s Duct utilizing basic dental tools, polyethylene tubing, and cyanoacrylate, and the appropriate rate of infusion. While this is the least traumatic method of delivery, the method is still limited by the volume able to be delivered (&#60;0.5 mL) and the potential for trauma to the duct and gland. We demonstrate using fluoroscopy that an infusate can be fully delivered into the gland, and further demonstrate by immunohistochemistry the transduction of a typical vector and expression of the delivered gene.

Salivary glands have been proposed as a tissue target site for gene therapy, especially in the area of vaccination by gene transfer. We demonstrate gene delivery in a non-human primate model utilizing retrograde parotid infusion.

ベクター化遺伝子導入のための非ヒト霊長類におけるステンセンダクトによる逆行性耳下腺注入

Salivary glands are an attractive tissue target for gene therapy with promising results already leading to human trials. They are inherently capable of ...

Radiation Treatment of Organotypic Cultures from Submandibular and Parotid Salivary Glands Models Key In Vivo Characteristics

Hyposalivation and xerostomia create chronic oral complications that decrease the quality of life in head and neck cancer patients who are treated with radiotherapy. Experimental approaches to understanding mechanisms of salivary gland dysfunction and restoration have focused on in vivo models, which are handicapped by an inability to systematically screen therapeutic candidates and efficiencies in transfection capability to manipulate specific genes. The purpose of this salivary gland organotypic culture protocol is to evaluate maximal time of culture viability and characterize cellular changes following ex vivo radiation treatment. We utilized immunofluorescent staining and confocal microscopy to determine when specific cell populations and markers are present during a 30-day culture period. In addition, cellular markers previously reported in in vivo radiation models are evaluated in cultures that are irradiated ex vivo. Moving forward, this method is an attractive platform for rapid ex vivo assessment of murine and human salivary gland tissue responses to therapeutic agents that improve salivary function.

Using three-dimensional organotypic cultures to visualize morphology and functional markers of salivary glands may provide novel insights into the mechanisms of tissue damage following radiation. Described here is a protocol to section, culture, irradiate, stain, and image 50&#8211;90 &#956;m thick salivary gland sections prior to and following exposure to ionizing radiation.

顎下および耳下腺唾液腺からの Organotypic 培養の放射線治療は生体内特性をモデル化する

Hyposalivation and xerostomia create chronic oral complications that decrease the quality of life in head and neck cancer patients who are treated with ...

マウス顎下腺再生のリアルタイムライブイメージング

Interrogating Cell-Cell Interactions in the Salivary Gland via Ex Vivo Live Cell Imaging

Salivary gland regeneration is a complex process involving intricate interactions among various cell types. Recent studies have shed light on the pivotal role played by macrophages in driving the regenerative response. However, our understanding of this critical role has primarily relied on static views obtained from fixed tissue biopsies. To overcome this limitation and gain insights into these interactions in real time, this study outlines a comprehensive protocol for culturing salivary gland tissue ex vivo and capturing live images of cell migration.
The protocol involves several key steps: First, mouse submandibular salivary gland tissue is carefully sliced using a vibratome and then cultured at an air-liquid interface. These slices can be intentionally injured, for instance, through exposure to radiation, to induce cellular damage and trigger the regenerative response. To track specific cells of interest, they can be endogenously labeled, such as by utilizing salivary gland tissue collected from genetically modified mice where a particular protein is marked with green fluorescent protein (GFP). Alternatively, fluorescently-conjugated antibodies can be employed to stain cells expressing specific cell surface markers of interest. Once prepared, the salivary gland slices are subjected to live imaging using a high-content confocal imaging system over a duration of 12 h, with images captured at 15 min intervals. The resulting images are then compiled to create a movie, which can subsequently be analyzed to extract valuable cell behavior parameters. This innovative method provides researchers with a powerful tool to investigate and better understand macrophage interactions within the salivary gland following injury, thereby advancing our knowledge of the regenerative processes at play in this dynamic biological context.

著者スポットライト:損傷後の唾液腺再生におけるマクロファージ-上皮細胞相互作用の研究 

Immunofluorescent imaging is constrained by the ability to observe complex, time-dependent biological processes in just a single snapshot in time. This study outlines a live-imaging approach conducted on precision-cut mouse submandibular gland slices. This approach allows for the real-time observation of cell-cell interactions during homeostasis and the processes of regeneration and repair.

唾液腺における細胞間相互作用の Ex Vivo Live Cell Imaging(英語)

Salivary gland regeneration is a complex process involving intricate interactions among various cell types. Recent studies have shed light on the pivotal ...