Name: application of hemostatic devices in laparoscopic hepatectomy
Uploaded: 2022-04-19T13:00:00
Description: Watch this Scientific Journal Video about Application of Hemostatic Devices in Laparoscopic Hepatectomy at JoVE.com

这项工作得到了广州市科技项目（202102010090）的资助。

该操作是例行的，并获得了道德认证。批准如下：广州医科大学附属第二医院临床研究与应用伦理委员会批准快速评审：科研伦理评审项目“简易易用止血装置在腹腔镜肝切除术中的应用”（受理号：2022-hg-ks-02）经广州医科大学附属第二医院临床研究应用伦理委员会批准。其研究内容和方法符合医学伦理规范和要求。
1. 术前准备、手术体位和麻醉
<ol><li>要求患...

<ol>
	<li>Hess, K. R., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Metastatic+patterns+in+adenocarcinoma.">Metastatic patterns in adenocarcinoma.</a> Cancer. 106 (7), 1624-1633 (2006).</li><li>van de Velde, C. J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=EURECCA+colorectal:+multidisciplinary+management:+European+consensus+conference+colon+&amp;+rectum.">EURECCA colorectal: multidisciplinary management: European consensus conference colon &amp; rectum.</a> European Journal of Cancer. 50 (1), 1-34 (2014).</li><li>Adams, R. B., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Selection+for+hepatic+resection+of+colorectal+liver+metastases:+expert+consensus+statement[J].">Selection for hepatic resection of colorectal liver metastases: expert consensus statement[J].</a> HPB. 15 (2), 91-103 (2013).</li><li>Reich, H., McGlynn, F., DeCaprio, J., Budin, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Laparoscopic+excision+of+benign+liver+lesions.">Laparoscopic excision of benign liver lesions.</a> Obstetrics and Gynecology. 78 (5), 956-958 (1991).</li><li>Ciria, R., Cherqui, D., Geller, D. A., Briceno, J., Wakabayashi, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Comparative+short-term+benefits+of+laparoscopic+liver+resection:+9000+cases+and+climbing.">Comparative short-term benefits of laparoscopic liver resection: 9000 cases and climbing.</a> Annals of Surgery. 263 (4), 761-777 (2016).</li><li>Makuuchi, M., Hasegawa, H., Yamazaki, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ultrasonically+guided+subsegmentectomy.">Ultrasonically guided subsegmentectomy.</a> Surgery, Gynecology, and Obstetrics. 161 (4), 346-350 (1985).</li><li>Agrawal, S., Belghiti, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Oncologic+resection+for+malignant+tumors+of+the+liver.">Oncologic resection for malignant tumors of the liver.</a> Annals of Surgery. 253 (4), 656-665 (2011).</li><li>Wakabayashi, T., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Laparoscopic+repeat+liver+resection+after+open+liver+resection:+A+comparative+study+from+a+single-centre.">Laparoscopic repeat liver resection after open liver resection: A comparative study from a single-centre.</a> Journal of Minimal Access Surgery. 16 (1), 59-65 (2020).</li><li>Berardi, G., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Parenchymal+sparing+anatomical+liver+resections+with+full+laparoscopic+approach:+description+of+technique+and+short-term+results.">Parenchymal sparing anatomical liver resections with full laparoscopic approach: description of technique and short-term results.</a> Annals of Surgery. 273 (4), 785-791 (2021).</li><li>Cho, J. Y., Han, H. S., Yoon, Y. S., Shin, S. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Feasibility+of+laparoscopic+liver+resection+for+tumors+located+in+the+posterosuperior+segments+of+the+liver,+with+a+special+reference+to+overcoming+current+limitations+on+tumor+location.">Feasibility of laparoscopic liver resection for tumors located in the posterosuperior segments of the liver, with a special reference to overcoming current limitations on tumor location.</a> Surgery. 144 (1), 32-38 (2008).</li><li>Ome, Y., Honda, G., Doi, M., Muto, J., Seyama, Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Laparoscopic+anatomic+liver+resection+of+segment+8+using+intrahepatic+Glissonean+approach.">Laparoscopic anatomic liver resection of segment 8 using intrahepatic Glissonean approach.</a> Journal of the American College of Surgeons. 230 (3), 13-20 (2020).</li><li>Ryu, T., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Perioperative+and+oncological+outcomes+of+laparoscopic+anatomical+hepatectomy+for+hepatocellular+carcinoma+introduced+gradually+in+a+single+center.">Perioperative and oncological outcomes of laparoscopic anatomical hepatectomy for hepatocellular carcinoma introduced gradually in a single center.</a> Surgical Endoscopy. 32 (2), 790-798 (2018).</li><li>Makabe, K., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Efficacy+of+occlusion+of+hepatic+artery+and+risk+of+carbon+dioxide+gas+embolism+during+laparoscopic+hepatectomy+in+a+pig+model.">Efficacy of occlusion of hepatic artery and risk of carbon dioxide gas embolism during laparoscopic hepatectomy in a pig model.</a> Journal of Hepato-Biliary-Pancreatic Sciences. 21 (8), 592-598 (2014).</li><li>Miyagi, S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Pure+laparoscopic+hepatectomy+combined+with+a+pure+laparoscopic+pringle+maneuver+in+patients+with+severe+cirrhosis.">Pure laparoscopic hepatectomy combined with a pure laparoscopic pringle maneuver in patients with severe cirrhosis.</a> Case Reports in Gastroenterology. 9 (1), 101-105 (2015).</li></ol>

肝切除术是原发性肝癌和继发性肝癌患者最重要的治疗方法之一。1991年，Reich等人应用腹腔镜切除肝脏良性肿瘤，并完成了世界上第一个腹腔镜肝切除术4。经过多年的快速发展和推广，腹腔镜肝切除术已成为治疗肝恶性肿瘤的主流方法之一，近年来得到了广泛的应用。该技术具有术中出血少、术后疼痛少、术后并发症发生率低、住院时间短等优点，该技术的可行性和安全性得到...

肝细胞癌是消化系统中最常见的恶性肿瘤之一。根据原发部位的不同，可分为原发性肝细胞癌和继发性肝癌。细胞可以从原发性肿瘤以各种方式从肝脏外的器官转移到肝脏，导致肝脏癌。据报道，肝脏中超过50%的转移细胞来自结直肠癌，而其他则来自乳腺，胰腺，肺和胃肿瘤等1。近年来，继发性肝癌的治疗方法很多，包括系统化疗、介入治疗、分子靶向治疗、手术等2.然而，根治性切除术仍然是最有效的治疗方法，因为它可以完全切除癌症3。
随着腹腔镜技术的飞速发展，腹腔镜解剖肝切除术逐渐被外科医生所认可，但仍未得到广泛应用，特别是在基层医疗机构。其中一个原因是需要高档止血装置。这些都是降低手术过程中出血和胆汁泄漏风险所必需的。在这里，我们提出了一套简单易行的止血设备，包括单腔导管，谐波手术刀和单极电凝术，用于进行腹腔镜肝切除术。为此，首先使用单腔导管间歇性地闭塞肝门。然后使用谐波手术刀切除肝实质组织。出血点通过单极电凝点对点凝。这种简单易行的止血设备使用单腔导管进行刺炙操作，并利用谐波手术刀和单极电凝术的止血。这些设备可以在医院轻松找到，从而为进行腹腔镜检查或培训提供了额外的便利。因此，这些简单易用的止血装置适用于在初级医疗机构中进行手术。
在这项研究中，患者是一名67岁的男性，被诊断患有肝脏中转移的乙状结肠中度分化腺癌。2021年1月进行了乙状结肠癌的根治性切除术。病理结果为中度分化腺癌，TNM分期为pT4aN2aM1。手术后给予四次FOLFOX化疗。在此之后，患者的身体状况被认为适合进行腹腔镜解剖肝切除术，以完全切除肝转移的病变。儿童-皮尤肝功能等级为A级。对于肝储备功能试验，ICG清除率试验中的R15为1.6%（<10%）。CT显示57 mm x 68 mm x 76 mm肿瘤横跨肝脏S5的背侧和S6的腹侧部分;三维重建模型如图 1所示。

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>HARMONIC ACE Ultrasonic Surgical Devices</td>
			<td>Ethicon Endo-Surgery,LLC</td>
			<td>HAR36</td>
			<td></td>
		</tr>
		<tr>
			<td>Hem-o-lock</td>
			<td>Teleflex Medical</td>
			<td>544233</td>
			<td></td>
		</tr>
		<tr>
			<td>Monopole Electrocoagulation</td>
			<td>KANGJI MEDICAL</td>
			<td>/</td>
			<td></td>
		</tr>
		<tr>
			<td>single lumen ureter</td>
			<td>WELL LEAD MEDICAL CO.,LTD.</td>
			<td>12F</td>
			<td></td>
		</tr>
	</tbody>
</table>

application of hemostatic devices in laparoscopic hepatectomy

腹腔镜肝切除术被认为是治疗良性和恶性肝脏疾病的常规方法，因为它是一种微创方法。尽管具有非侵入性方面，但出血和胆汁渗漏发生在手术期间或术后肝实质组织切除术中，表明需要高级别止血装置，如超声手术抽吸，双极电凝等。这些高级止血装置的可用性不足，阻止了腹腔镜肝切除术成为基础医疗机构的普遍手术。针对上述情况，该方案开发了一套简单易行的止血装置，包括谐波手术刀、单极电凝和单腔导管，以创新方式进行肝实质组织切除术。首先，用单腔导管间歇性地闭塞肝门或肝椎弓根，然后夹紧15分钟并释放5分钟。随后，使用谐波手术刀，对肝脏进行钳夹和粉碎，以切断肝实质组织并揭示肝内动脉，静脉和胆管。最后，在每个点使用单极电凝来凝固出血点。利用这些方法可见肝内管路结构，可容易止血，降低胆汁渗漏发生率，提高腹腔镜肝切除术的安全性和可行性。因此，这里显示的简单易用的止血装置适用于初级医疗机构进行手术。

手术在2.5 h内完成，术中出血量为100 mL，未输血。无短期并发症，术后第8 天出院。操作后癌胚抗原（CEA）水平从操作前的1058.69μg/L降至110.64μg/L（图12）。手术后两个月，患者返回医院继续化疗（化疗方案为mFOLFOX6）。根据肿瘤病理结果进行化疗方案，如图 11所示。将患者返回医院复查时进行的CT检查与手术前进行的CT检查进行比较。没有观察到...

Watch this Scientific Journal Video about Application of Hemostatic Devices in Laparoscopic Hepatectomy at JoVE.com

Application of Hemostatic Devices in Laparoscopic Hepatectomy

高级止血装置对于腹腔镜肝切除术至关重要。然而，这些设备在基础医疗组织中并不普遍。因此，本文展示了一套简单易行的止血装置，可以使腹腔镜肝切除术更容易进行。

止血装置在腹腔镜肝切除术中的应用

Laparoscopic hepatectomy is considered a conventional method for treating benign and malignant liver diseases because it is a minimally invasive method. ...

该技术的主要优点是止血装置简单易获得，可以在基础医院进行。这套止血设备可进一步应用于腹腔镜肝切除术等系统。强烈建议有一定的腹腔镜手术基础，特别是熟练使用谐波手术刀将这套止血装置应用于腹腔镜肝切除术。首先在腹腔镜检查中应用直接穿刺以建立气腹。在脐下区域，肋缘下方的右锁骨中线和腹侧中线的每个观察孔中放置一个10毫米的穿刺器。然后将一个五毫米的穿刺器放在右前辅助线和腹侧中线。从谐波手术刀、单极电凝和单腔导管开始组装止血装置，以创新方式用于肝实质组织切除术。要执行普林格尔操作，请使用单腔导管间歇性地闭塞第一个肝门。用止血夹夹住腔导管15分钟，然后使用谐波手术刀将其松开5分钟，切断夹子并将夹子取出以恢复第一个肝门的血液供应。分离供应肿瘤的肝椎弓根。用谐波手术刀将它们逐个切除。观察肝表面的缺血线，在肝蒂切除后变得可见。使用钩子标记它。使用谐波手术刀切除肝实质组织，同时逐渐夹紧和压碎肝脏。观察肝内动脉、静脉和胆管。切除和分割这些管线结构。一旦出现出血，立即使用点对点单极电凝术凝固肝清扫表面上的出血点。使用右肝后蒂作为肝内实质组织切除术的路标。保留主要的肝后椎弓根并解剖供应肿瘤的椎弓根分支。要完成手术，请清洗肝脏表面，以便查看是否仍有出血点。止肝清扫表面出血，从腹腔中取出肿瘤标本。操作在2.5小时内完成。术中出血量为100毫升，未输血，未观察到短期并发症。术后癌胚抗原水平从术前每升1058.69微克降至每升110.64微克。根据肿瘤病理结果，手术后两个月恢复化疗治疗。术后CT检查与术前CT比较，未观察到肿瘤再生迹象。手术过程中最重要的一步是使用谐波手术刀。需要大量的技能来避免对重要结构的损坏。

application-of-hemostatic-devices-in-laparoscopic-hepatectomy

Research

JoVE Journal

Cancer Research

Evaporation-reducing Culture Condition Increases the Reproducibility of Multicellular Spheroid Formation in Microtiter Plates

Tumor models that closely imitate in vivo conditions are becoming increasingly popular in drug discovery and development for the screening of potential anti-cancer drugs. Multicellular tumor spheroids (MCTSes) effectively mimic the physiological conditions of solid tumors, making them excellent in vitro models for lead optimization and target validation. Out of the various techniques available for MCTS culture, the liquid-overlay method on agarose is one of the most inexpensive methods for MCTS generation. However, the reliable transfer of MCTS cultures using liquid-overlay for high-throughput screening may be compromised by a number of limitations, including the coating of microtiter plates (MPs) with agarose and the irreproducibility of uniform MCTS formation across wells. MPs are significantly prone to edge effects that result from excessive evaporation of medium from the exterior of the plate, preventing the use of the entire plate for drug tests. This manuscript provides detailed technical improvements to the liquid-overlay technique to increase the scalability and reproducibility of uniform MCTS formation. Additionally, details on a simple, semi-automatic, and universally applicable software tool for the evaluation of MCTS features after drug treatment is presented.

The uneven loss of medium from microtiter plates affects the reproducibility of uniform multicellular tumor spheroid formation. Improving culture conditions to reduce significant medium loss will improve the reproducibility of spheroid formation and the results of spheroid-based assays using the liquid-overlay technique.

蒸发减少培养条件的提高多细胞球体形成的微孔板重现

Tumor models that closely imitate in vivo conditions are becoming increasingly popular in drug discovery and development for the screening of potential ...

科研

癌症研究

Application of Laser Micro-irradiation for Examination of Single and Double Strand Break Repair in Mammalian Cells

Highly coordinated DNA repair pathways exist to detect, excise and replace damaged DNA bases, and coordinate repair of DNA strand breaks. While molecular biology techniques have clarified structure, enzymatic functions, and kinetics of repair proteins, there is still a need to understand how repair is coordinated within the nucleus. Laser micro-irradiation offers a powerful tool for inducing DNA damage and monitoring the recruitment of repair proteins. Induction of DNA damage by laser micro-irradiation can occur with a range of wavelengths, and users can reliably induce single strand breaks, base lesions and double strand breaks with a range of doses. Here, laser micro-irradiation is used to examine repair of single and double strand breaks induced by two common confocal laser wavelengths, 355 nm and 405 nm. Further, proper characterization of the applied laser dose for inducing specific damage mixtures is described, so users can reproducibly perform laser micro-irradiation data acquisition and analysis.

Confocal fluorescence microscopy and laser micro-irradiation offer tools for inducing DNA damage and monitoring the response of DNA repair proteins in selected sub-nuclear areas. This technique has significantly advanced our knowledge of damage detection, signaling, and recruitment. This manuscript demonstrates these technologies to examine single and double strand break repair.

激光微辐照在哺乳动物细胞单、双链断裂修复中的应用

Highly coordinated DNA repair pathways exist to detect, excise and replace damaged DNA bases, and coordinate repair of DNA strand breaks. While molecular ...

Laparoscopic Pancreatoduodenectomy With Modified Blumgart Pancreaticojejunostomy

Minimally invasive pancreatic resections are technically demanding but rapidly increasing in popularity. In contrast to laparoscopic distal pancreatectomy, laparoscopic pancreatoduodenectomy (LPD) has not yet obtained wide acceptance, probably due to technical challenges, especially regarding the pancreatic anastomosis.
The study describes and demonstrates all steps of LPD, including the modified Blumgart pancreaticojejunostomy. Indications for LPD are all pancreatic and peri-ampullary tumors without vascular involvement. Relative contra-indications are body mass index &#62;35 kg/m2, chronic pancreatitis, mid-cholangiocarcinomas and large duodenal cancers.
The patient is in French position, 6 trocars are placed, and dissection is performed using an (articulating) sealing device. A modified Blumgart end-to-side pancreaticojejunostomy is performed with 4 large needles (3/0) barbed trans-pancreatic sutures and 4 to 6 duct-to-mucosa sutures using 5/0 absorbable multifilament combined with a 12 cm, 6 or 8 Fr internal stent using 3D laparoscopy. Two surgical drains are placed alongside the pancreaticojejunostomy.
The described technique for LPD including a modified Blumgart pancreatico-jejunostomy is well standardized, and its merits are currently studied in the randomized controlled multicenter trial. This complex operation should be performed at high-volume centers where surgeons have extensive experience in both open pancreatic surgery and advanced laparoscopic gastro-intestinal surgery.

Laparoscopic pancreatoduodenctomy (LPD) may offer advantages over open pancreatoduodenectomy, including early postoperative mobilization, less delayed gastric emptying and a shorter hospital stay. However, LPD is technically challenging and not well-standardized, especially regarding the pancreatic anastomosis. We describe a standardized technique for the pancreatic anastomosis during LPD: modified Blumgart pancreaticojejunostomy.

改良 Blumgart Pancreaticojejunostomy 腹腔镜胰十二指肠切除术

Minimally invasive pancreatic resections are technically demanding but rapidly increasing in popularity. In contrast to laparoscopic distal ...

The Application of 1% Methylene Blue Dye As a Single Technique in Breast Cancer Sentinel Node Biopsy

In this study, we injected 1% methylene blue dye (MBD) into the subareolar or peritumoral space of the breast. In the case of breast conserving surgery (BCS), a separate incision in the lower axilla hairline was made to find the sentinel nodes (SNs). In mastectomy, the SNs were identified through the same mastectomy incision. The SNs were described as blue nodes or nodes with lymphatic blue channels. An anatomical landmark in the axilla was used to facilitate SNs identification. The SNs metastases were evaluated by intraoperative frozen section analysis and histopathology examination as it is a gold standard. Here, we described the MBD as the lone technique in breast cancer sentinel node biopsy (SNB) which could be useful when radioisotope tracer or patent or isosulfan blue dye (PBD) cannot be provided.

In Indonesia, sentinel node biopsy (SNB) is not routinely performed for breast cancer surgery because of the limitation to provide radioisotope tracer and isosulfan or patent blue dye (PBD). To overcome this obstacle, we applied 1% methylene blue dye (MBD) as a single agent to map the sentinel nodes (SNs).

1%亚甲蓝染料作为单一技术在乳腺癌哨点节点活检中的应用

In this study, we injected 1% methylene blue dye (MBD) into the subareolar or peritumoral space of the breast. In the case of breast conserving surgery ...

The Clinical Application of Tumor Treating Fields Therapy in Glioblastoma

Glioblastoma is the most common and lethal form of brain cancer, with a median survival of 15 months after diagnosis and a 5 year survival rate of only 5% with current standard of care. Tumors often recur within 9 months following initial surgery, radiation and chemotherapy, at which point treatment options become limited. This highlights the pressing need for the development of better therapeutics to prolong survival and increase the quality of life for these patients.
Tumor Treating Fields (TTFields) therapy was developed to take advantage of the effect of low frequency alternating electrical fields on cells for cancer therapy. TTFields have been demonstrated to disrupt cells during mitosis and slow tumor growth. There is also growing evidence that they act through stimulating immune responses within exposed tumors. The advantages of TTFields therapy include its noninvasive approach and increased quality of life compared to other treatment modalities such as cytotoxic chemotherapies. The Food and Drug Administration approved TTFields therapy for the treatment of recurrent glioblastoma in 2011 and for newly diagnosed glioblastoma in 2015. We report on the effects of TTFields during mitosis, the results of electric fields modeling, and proper transducer array placement. Our protocol outlines the clinical application of TTFields on a patient post-surgery, using the second-generation device.

Glioblastoma is the most common and aggressive primary brain malignancy in adults, with most tumors recurring after initial treatment. Tumor Treating Fields (TTFields) therapy is the newest treatment modality for glioblastoma. Here, we describe the proper application of TTFields-transducer arrays on patients and discuss theory and aspects of treatment.

肿瘤治疗胶质母细胞瘤的临床应用

Glioblastoma is the most common and lethal form of brain cancer, with a median survival of 15 months after diagnosis and a 5 year survival rate of only 5% ...

In Vitro Evaluation of Oncogenic Transformation in Human Mammary Epithelial Cells

Tumorigenesis is a multi-step process in which cells acquire capabilities&#160;that allow their growth, survival, and dissemination under hostile conditions. Different tests seek to identify and quantify these hallmarks of cancerous cells; however, they often focus on a single aspect of cellular transformation and, in fact, multiple tests are required for their proper characterization. The purpose of this work is to provide researchers with a set of tools to assess cellular transformation in vitro from a broad perspective, thereby making it possible to draw sound conclusions.
A sustained proliferative signaling activation is the major feature of tumoral tissues and can be easily monitored under in vitro conditions by calculating the number of population doublings achieved over time. Besides, the growth of cells in 3D cultures allows their interaction with surrounding cells, resembling what occurs in vivo. This enables the evaluation of cellular aggregation and, together with immunofluorescent labeling of distinctive cellular markers, to obtain information on another relevant feature of tumoral transformation: the loss of proper organization. Another remarkable characteristic of transformed cells is their capacity to grow without attachment to other cells and to the extracellular matrix, which can be evaluated with the anchorage assay.
Detailed experimental procedures to evaluate cell growth rate, to perform immunofluorescent labeling of cell lineage markers in 3D cultures, and to test anchorage-independent cell growth in soft agar are provided. These methodologies are optimized for Breast Primary Epithelial Cells (BPEC) due to its relevance in breast cancer; however, procedures can be applied to other cell types after some adjustments.

This protocol provides experimental in vitro tools to evaluate the transformation of human mammary cells. Detailed steps to follow-up cell proliferation rate, anchorage-independent growth capacity, and distribution of cell lineages in 3D cultures with basement membrane matrix are described.

人类乳腺上皮细胞致癌转化的体外评价

Tumorigenesis is a multi-step process in which cells acquire capabilities&#160;that allow their growth, survival, and dissemination under hostile ...

Co-culture of Glutamatergic Neurons and Pediatric High-Grade Glioma Cells Into Microfluidic Devices to Assess Electrical Interactions

Pediatric high-grade gliomas (pHGG) represent childhood and adolescent brain cancers that carry a rapid dismal prognosis. Since there is a need to overcome the resistance to current treatments and find a new way of cure, modeling the disease as close as possible in an in vitro setting to test new drugs and therapeutic procedures is highly demanding. Studying their fundamental pathobiological processes, including glutamatergic neuron hyperexcitability, will be a real advance in understanding interactions between the environmental brain and pHGG cells. Therefore, to recreate neurons/pHGG cell interactions, this work shows the development of a functional in vitro model co-culturing human-induced Pluripotent Stem (hiPS)-derived cortical glutamatergic neurons pHGG cells into compartmentalized microfluidic devices and a process to record their electrophysiological modifications. The first step was to differentiate and characterize human glutamatergic neurons. Secondly, the cells were cultured in microfluidic devices with pHGG derived cell lines. Brain microenvironment and neuronal activity were then included in this model to analyze the electrical impact of pHGG cells on these micro-environmental neurons. Electrophysiological recordings are coupled using multielectrode arrays (MEA) to these microfluidic devices to mimic physiological conditions and to record the electrical activity of the entire neural network. A significant increase in neuron excitability was underlined in the presence of tumor cells.

Recent works uncover the neuronal impact on high-grade pediatric glioma (pHGG) cells and their reciprocal interactions. The present work shows the development of an in vitro model co-culturing pHGG cells and glutamatergic neurons and recorded their electrophysiological interactions to mimic those interactivities.

将谷氨酸能神经元和儿科高级别胶质瘤细胞共培养到微流体装置中以评估电相互作用

Pediatric high-grade gliomas (pHGG) represent childhood and adolescent brain cancers that carry a rapid dismal prognosis. Since there is a need to ...

Flexible Organic Electronic Devices for Pulsed Electric Field Therapy of Glioblastoma

Glioblastoma is difficult to eradicate with standard oncology therapies due to its high degree of invasiveness. Bioelectric treatments based on pulsed electric fields (PEFs) are promising for the improvement of treatment efficiency. However, they rely on rigid electrodes that cause acute and chronic damage, especially in soft tissues such as the brain. In this work, flexible electronics were used to deliver PEFs to tumors and the biological response was evaluated with fluorescent microscopy. Interdigitated gold electrodes on a thin, transparent parylene-C substrate were coated with the conducting polymer PEDOT:PSS, resulting in a conformable and biocompatible device. The effects of PEFs on tumors and their microenvironment were examined using various biological models. First, monolayers of glioblastoma cells were cultured on top of the electrodes to investigate phenomena in vitro. As an intermediate step, an in ovo model was developed where engineered tumor spheroids were grafted in the embryonic membrane of a quail. Due to the absence of an immune system, this led to highly vascularized tumors. At this early stage of development, embryos have no immune system, and tumors are not recognized as foreign bodies. Thus, they can develop fast while developing their own vessels from the existing embryo vascular system, which represents a valuable 3D cancer model. Finally, flexible electrode delivery of PEFs was evaluated in a complete organism with a functional immune system, using a syngenic, orthograft (intracranial) mouse model. Tumor spheroids were grafted into the brain of transgenic multi-fluorescent mice prior to the implantation of flexible organic electrode devices. A sealed cranial window enabled multiphoton imaging of the tumor and its microenvironment during treatment with PEFs over a period of several weeks.

This work describes the development of flexible interdigitated electrodes for implementation in 3D brain tumor models, namely, in vitro culture, in ovo model, and in vivo murine model. The proposed method can be used to evaluate the effects of pulsed electric fields on tumors at different levels of complexity.

用于胶质母细胞瘤脉冲电场治疗的柔性有机电子器件

Glioblastoma is difficult to eradicate with standard oncology therapies due to its high degree of invasiveness. Bioelectric treatments based on pulsed ...

Generation and Culturing of High-Grade Serous Ovarian Cancer Patient-Derived Organoids

Organoids are 3D dynamic tumor models that can be grown successfully from patient-derived ovarian tumor tissue, ascites, or pleural fluid and aid in the discovery of novel therapeutics and predictive biomarkers for ovarian cancer. These models recapitulate clonal heterogeneity, the tumor microenvironment, and cell-cell and cell-matrix interactions. Additionally, they have been shown to match the primary tumor morphologically, cytologically, immunohistochemically, and genetically. Thus, organoids facilitate research on tumor cells and the tumor microenvironment and are superior to cell lines. The present protocol describes distinct methods to generate patient-derived ovarian cancer organoids from patient tumors, ascites, and pleural fluid samples with a higher than 97% success rate. The patient samples are separated into cellular suspensions by both mechanical and enzymatic digestion. The cells are then plated utilizing a basement membrane extract (BME) and are supported with optimized growth media containing supplements specific to the culturing of high-grade serous ovarian cancer (HGSOC). After forming initial organoids, the PDOs can sustain long-term culture, including passaging for expansion for subsequent experiments.

Patient-derived organoids (PDO) are a three-dimensional (3D) culture that can mimic the tumor environment in vitro. In high-grade serous ovarian cancer, PDOs represent a model to study novel biomarkers and therapeutics.

高级别浆液性卵巢癌患者来源类器官的生成和培养

Organoids are 3D dynamic tumor models that can be grown successfully from patient-derived ovarian tumor tissue, ascites, or pleural fluid and aid in the ...

Application of the En Bloc Concept Combined with Anatomic Resection in Laparoscopic Hepatectomy

Laparoscopic hepatectomy has been reported in many studies, and it is the mainstream method of liver resection. In some particular cases, such as when there are tumors adjacent to the cystic bed, surgeons cannot palpate the surgical margins through the laparoscopic approach, which leads to uncertainty about R0 resection. Conventionally, the gallbladder is resected first, and the hepatic lobes or segments are resected second. However, tumor tissues can be disseminated in the above cases. To address this issue, based on the recognition of the porta hepatis and intrahepatic anatomy, we propose a unique approach to hepatectomy combined with gallbladder resection by en bloc anatomic resection in situ. Firstly, after dissecting the cystic duct, without cutting the gallbladder primarily, the porta hepatis is pre-occluded by the single lumen ureter; secondly, the left hepatic pedicle is made free by the gap of the Laennec membrane and Hilar plate; thirdly, the assistant is asked to drag the fundus of the gallbladder, and the liver parenchyma tissue is resected using a harmonic scalpel along the ischemia line on the liver surface and intraoperative ultrasound. The whole middle hepatic vein (MHV) and its tributaries appear completely; lastly, the left hepatic vein (LHV) is disconnected, and the specimen is taken out from the abdominal cavity. The tumor, gallbladder, and other surrounding tissues are resected en bloc, which meets the tumor-free criterion, and a wide incisal margin and R0 resection are achieved. Therefore, the laparoscopic hepatectomy with the combination of the en bloc concept and anatomic resection is a safe, effective, and radical method with low postoperative recurrence and metastasis.

Numerous studies have demonstrated the advantages of anatomic resection. Nonetheless, whether anatomic resection can increase R0 resection rates remains controversial. Consequently, the present study describes an innovative procedure involving the en bloc concept combined with anatomic resection in laparoscopic hepatectomy, which can reduce postoperative recurrence and metastasis.

整体概念联合解剖切除在腹腔镜肝切除术中的应用

Laparoscopic hepatectomy has been reported in many studies, and it is the mainstream method of liver resection. In some particular cases, such as when ...