The authors have nothing to acknowledge.

Tutte le procedure sono stati rivisti e approvati dal Institutional Review Board della Chungnam National University Hospital. 1. Misura con ecografia <ol><li> Impostare la postura del paziente. <ol><li> Posizionare il braccio del paziente in posizione supina, con entrambe le braccia sul letto. </li></ol></li><li> Determinare i siti di misurazione sulla parte superiore del paziente. <ol><li> Mark nel punto centrale della piega del polso, il punto intermedio tra il mediale e epicondili laterali a livello del gomito, e la scanalatura bicipitale da una penna. </li><li> Utilizzando una penna collegare direttamente questi tre punti da un righello. Segnare le due aree bersaglio sulla parte superiore del braccio e dell&#39;avambraccio (Figura 1). <ol><li> Segnare la parte prossimale, 10 centimetri in prossimità del punto di piega lungo la linea tra il gomito e solco bicipitale. </li><li> Segnare la parte distale 10 cm distalmente al punto di piega lungo la linea tra il gomito e polso. </li></ol></li></ol></li><li> Misurare lo spessore del tessuto sottocutaneo e muscolare degli arti superiori. La procedura deve essere eseguita da un ecografista esperto. <ol><li> Accendere il sistema a ultrasuoni. Inserire modalità 2D dei tessuti molli tramite la tastiera. Impostare la profondità di 5 cm. <ol><li> Scegliere il trasduttore lineare 14L5. </li><li> Applicare gel sufficienti al trasduttore ad ultrasuoni. Focus sul tessuto sottocutaneo. </li></ol></li><li> Posizionare il trasduttore ad ultrasuoni perpendicolarmente all&#39;asse ventrale arto superiore. Utilizzare l&#39;asse corto. Acquisire l&#39;immagine in cui lo spessore del gel è di almeno 1 cm e il contorno tessuto molle non è distorta. </li><li> Misurare le spessori del muscolo e del tessuto sottocutaneo. <ol><li> Disegna una linea selezionando il calibratore nel software tramite la tastiera del sistema ad ultrasuoni </li><li> Misurare lo spessore del muscolo, definita come la distanza dal punto più alto sul bordo dell&#39;osso al punto più altosulla porzione di confine della fascia (Figura 2). </li><li> Misurare lo spessore del tessuto sottocutaneo, definita come la distanza dalla pelle alla fascia. </li></ol></li></ol></li></ol> 2. Applicazione clinica di Ecografia a BCRL <ol><li> Misurare lo spessore del tessuto sottocutaneo e muscolare dell&#39;arto superiore al basale. <ol><li> Eseguire le misurazioni di base nell&#39;ordine indicato al punto 1. </li></ol></li><li> Chiedere al paziente di eseguire una serie di PRE con un 0,5 kg manubrio 14. <ol><li> Dare al paziente una calza di compressione o un bendaggio multistrato da indossare. Scegliere un 0,5 kg manubrio. </li><li> Chiedere al paziente di eseguire le PRE nel seguente ordine: (1) manubri vola, (2) tricipite estensione, (3) un braccio fila piegato-over, (4) bicipiti curl, (5) rilancio lato manubrio, e (6 ) sollevando le braccia in avanti (figura 3). <ol><li> Al paziente di completare la 6esercizi due volte al giorno, secondo il seguente calendario: 5 volte ciascuno durante la settimana 1 °, per 10 volte durante la settimana 2 °, 15 volte ciascuno durante la settimana 3 °, 20 volte ciascuno durante la 4 ° settimana, e 25 volte ciascuno durante 5 settimane th -8 ° (Figura 4). </li></ol></li></ol></li><li> Utilizzare ecografia per identificare gli effetti terapeutici della PRE rispetto alla terapia convenzionale. <ol><li> Misurare lo spessore del tessuto sottocutaneo e muscolare dell&#39;arto superiore a 4 e 8 settimane. Eseguire le misurazioni nell&#39;ordine indicato al punto 1. </li></ol></li><li> Calcolare lo spessore del tessuto sottocutaneo e muscolare al basale, 4 settimane e 8 settimane. </li><li> Analizzare i dati. <ol><li> Confrontare la differenza tra i due bracci con un t-test accoppiato. Confronta i cambiamenti nel tessuto sottocutaneo e dello spessore del muscolo utilizzando un ripetute-misure ANOVA. </li></ol></li></ol>

<ol>
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H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Does+the+effect+of+weight+lifting+on+lymphedema+following+breast+cancer+differ+by+diagnostic+method:+results+from+a+randomized+controlled+trial.">Does the effect of weight lifting on lymphedema following breast cancer differ by diagnostic method: results from a randomized controlled trial.</a> Breast Cancer Res Treat. 130 (1), 227-234 (2011).</li><li>Fu, M. R., Ridner, S. H., Armer, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Post-breast+cancer+lymphedema:+part+1.">Post-breast cancer lymphedema: part 1.</a> Am J Nurs. 109 (7), 48-54 (2009).</li><li>Rockson, S. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ultrasonography+in+the+Evaluation+of+Breast+Cancer-Related+Lymphedema.">Ultrasonography in the Evaluation of Breast Cancer-Related Lymphedema.</a> Lymphatic Research and Biology. 14 (1), 1 (2016).</li><li>Suehiro, 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=Significance+of+Ultrasound+Examination+of+Skin+and+Subcutaneous+Tissue+in+Secondary+Lower+Extremity+Lymphedema.">Significance of Ultrasound Examination of Skin and Subcutaneous Tissue in Secondary Lower Extremity Lymphedema.</a> Ann Vasc Dis. 6 (2), 180-188 (2013).</li><li>Lee, J. H., 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=Ultrasonographic+Evaluation+of+Therapeutic+Effects+of+Complex+Decongestive+Therapy+in+Breast+Cancer-Related+Lymphedema.">Ultrasonographic Evaluation of Therapeutic Effects of Complex Decongestive Therapy in Breast Cancer-Related Lymphedema.</a> Ann Rehabil Med. 37 (5), 683-689 (2013).</li><li>DiSipio, T., Rye, S., Newman, B., Hayes, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Incidence+of+unilateral+arm+lymphoedema+after+breast.">Incidence of unilateral arm lymphoedema after breast.</a> Lancet Oncol. 14 (6), 500-515 (2013).</li><li>Stanton, A. W., 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=Lymphatic+drainage+in+the+muscle+and+subcutis+of+the+arm+after+breast+cancer+treatment.">Lymphatic drainage in the muscle and subcutis of the arm after breast cancer treatment.</a> Breast Cancer Res Treat. 117 (3), 549-557 (2008).</li><li>Godoy, M. F. G., Pereira, M. R., Oliani, A. H., Godoy, J. M. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Synergic+Effect+of+Compression+Therapy+and+Controlled+Active+Exercises+Using+a+Facilitating+Device+in+the+Treatment+of+Arm+Lymphedema.">Synergic Effect of Compression Therapy and Controlled Active Exercises Using a Facilitating Device in the Treatment of Arm Lymphedema.</a> Int J Med Sci. 9 (4), 280-284 (2012).</li><li>Cormie, P., Pumpa, K., Galv&#227;o, D. A., Turner, E., Spry, N., Saunders, C., Zissiadis, Y., Newton, R. U. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Is+it+safe+and+efficacious+for+women+with+lymphedema+secondary+to+breast+cancer+to+lift+heavy+weights+during+exercise:+a+randomised+controlled+trial.">Is it safe and efficacious for women with lymphedema secondary to breast cancer to lift heavy weights during exercise: a randomised controlled trial.</a> J Cancer Surviv. 7 (3), 413-424 (2013).</li><li>Bok, S. K., Jeon, Y., Hwang, P. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ultrasonographic+evaluation+of+the+effects+of+progressive+resistive+exercise+in+breast+cancer-related+lymphedema.">Ultrasonographic evaluation of the effects of progressive resistive exercise in breast cancer-related lymphedema.</a> Lymphatic Research and Biology. 14 (1), 18-24 (2016).</li><li>Milne, H. M., Wallman, K. E., Gordon, S., Courneya, K. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effects+of+a+combined+aerobic+and+resistance+exercise+program+in+breast+cancer+survivors:+a+randomized+controlled+trial.">Effects of a combined aerobic and resistance exercise program in breast cancer survivors: a randomized controlled trial.</a> Breast Cancer Res Treat. 108 (2), 279-288 (2008).</li><li>Schmitz, K. H., 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=Weight+lifting+for+women+at+risk+for+breast-cancer-related+lymphedema.">Weight lifting for women at risk for breast-cancer-related lymphedema.</a> JAMA. 304 (24), 2699-2705 (2010).</li><li>Kiel, K. D., Rademacker, A. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Early-stage+breast+cancer:+Arm+edema+after+wide+excision+and+breast+irradiation.">Early-stage breast cancer: Arm edema after wide excision and breast irradiation.</a> Radiology. (1), 279-283 (1996).</li></ol>

The authors have nothing to disclose.

Le valutazioni di BCRL possono essere condotte attraverso varie tecniche, tra cui circonferenze del braccio di misurazione del nastro, misura dello spostamento d&#39;acqua, la spettroscopia bioimpedenza, perometry (volumetro optoelettronica), linfoscintigrafia, la diagnosi medico, o gonfiore self-report da parte del paziente. Tuttavia, non esistono standard per la misurazione della BCRL; la diagnosi e la durata del trattamento può essere determinato empiricamente in vari modi, a seconda della clinica. <p class="jov...

Linfedema è definita come una condizione di localizzato ricca di proteine ritenzione idrica e gonfiore dei tessuti causate da un 1,2 sistema linfatico compromessa. Come linfedema progredisce, fibrociti e / o adipociti proliferano nelle zone colpite, che porta a cambiamenti nella struttura della pelle e del tessuto sottocutaneo e una maggiore vulnerabilità alle infezioni batteriche e fungine 3,4. L&#39;osservazione di questi cambiamenti in varie parti degli arti può chiarire ulteriormente la gravità e l&#39;estensione del linfedema. Il sistema linfatico sottofasciale contribuisce alla gravità del linfedema più del sistema linfatico epifasciale. Recenti ricerche hanno riguardato principalmente l&#39;uso dell&#39;ecografia nella diagnosi di linfedema 5,6. L&#39;ecografia è un metodo relativamente poco costoso per osservare le caratteristiche dei tessuti molli. Un precedente studio ha riportato che linfedema può essere attendibilmente diagnosticato con ecografia confrontando tha spessore del tessuto cutaneo e sottocutaneo e anche valutando la conformità del tessuto sottocutaneo in un ambiente 7,8 clinica. Inoltre, ecografia è ampiamente usato in clinica per la sua facile accessibilità, di sicurezza da esposizione a radiazioni, ed economicità 5,6. A dispetto di questi benefici, gli studi sulla ecografia per linfedema solo sono stati concentrati sulla diagnosi, non sulla valutazione degli interventi terapeutici. Pertanto, lo studio mirato a determinare non solo l&#39;utilità dell&#39;ecografia nel chiarire gli effetti di PRE sulla BCRL, ma anche se ultrasonografia ha un significato clinico come test di follow-up nel confermare l&#39;efficacia del trattamento. Questo studio è stato condotto per stabilire il protocollo per ultrasuoni eseguiti per esaminare linfedema.

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ultrasonographic evaluation of breast cancer-related lymphedema

Linfedema è una delle più comuni complicanze dopo chirurgia del cancro al seno. Ci sono molti strumenti diagnostici per linfedema, ma nessun metodo standard ancora esiste. Progressive Resistenza Esercizio (PRE) dovrebbe migliorare linfedema senza gonfiore aggiuntivo. Questo studio ha mostrato effetti terapeutici di PRE sul linfedema mediante ecografia per misurare la variazione di spessore del muscolo e del tessuto sottocutaneo. Lo spessore del tessuto sottocutaneo è diminuito di più nel gruppo PRE che nel gruppo non-PRE. L&#39;ecografia è ampiamente usato in molte cliniche per la sua facile accessibilità, la sicurezza, ed economicità. L&#39;ecografia è uno dei migliori strumenti per la diagnosi e la determinazione l&#39;efficacia del trattamento sul linfedema al seno per cancro (BCRL).

Questo studio ha incluso 32 pazienti con diagnosi di BCRL. Lo spessore del muscolo e del tessuto sottocutaneo e le circonferenze degli arti superiori prossimale e distale sono stati misurati al basale, 4 settimane e 8 settimane dopo PRE. Esaminatori misurato la circonferenza degli arti superiori con un nastro misura, e quindi lo spessore del muscolo e del tessuto sottocutaneo sono stati misurati mediante ecografia. I pazienti sono stati divisi casualmente in un gruppo PRE e un gruppo non...

Watch this Scientific Journal Video about Ultrasonographic Evaluation of Breast Cancer-related Lymphedema at JoVE.com

Ultrasonographic Evaluation of Breast Cancer-related Lymphedema

Questo studio ha utilizzato l&#39;ecografia per la diagnosi e test di follow-up per confermare l&#39;efficacia del trattamento di progressiva resistenza esercizio (PRE) in linfedema al seno per cancro (BCRL). L&#39;ecografia può essere applicato in modo efficace per garantire la diagnosi e il trattamento del linfedema.

Ecografica Valutazione del seno Linfedema correlati al cancro

Lymphedema is one of the most common complications after breast cancer surgery. There are many diagnostic tools for lymphedema, but no standard method yet ...

ultrasonographic-evaluation-of-breast-cancer-related-lymphedema

Research

JoVE Journal

Cancer Research

9.1K Views.  Chungnam National University Hospital (CNUH).  Questo studio ha utilizzato l'ecografia per la diagnosi e test di follow-up per confermare l'efficacia del trattamento di progressiva resistenza esercizio (PRE) in linfedema al seno per cancro (BCRL). L'ecografia può essere applicato in modo efficace per garantire la diagnosi e il trattamento del linfedema. 

Video: Ultrasonographic Evaluation of Breast Cancer-related Lymphedema

Utilizing Functional Genomics Screening to Identify Potentially Novel Drug Targets in Cancer Cell Spheroid Cultures

The identification of functional driver events in cancer is central to furthering our understanding of cancer biology and indispensable for the discovery of the next generation of novel drug targets. It is becoming apparent that more complex models of cancer are required to fully appreciate the contributing factors that drive tumorigenesis in vivo and increase the efficacy of novel therapies that make the transition from pre-clinical models to clinical trials.
Here we present a methodology for generating uniform and reproducible tumor spheroids that can be subjected to siRNA functional screening. These spheroids display many characteristics that are found in solid tumors that are not present in traditional two-dimension culture. We show that several commonly used breast cancer cell lines are amenable to this protocol. Furthermore, we provide proof-of-principle data utilizing the breast cancer cell line BT474, confirming their dependency on amplification of the epidermal growth factor receptor HER2 and mutation of phosphatidylinositol-4,5-biphosphate 3-kinase (PIK3CA) when grown as tumor spheroids. Finally, we are able to further investigate and confirm the spatial impact of these dependencies using immunohistochemistry.

Identifying novel drug targets that transition from pre-clinical testing to human trials is a scientific priority. To that end, here we describe a functional genomics approach for examining the impact of gene depletion on cancer cell line spheroids, which more appropriately model human cancers in vivo.

Utilizzando Genomica Funzionale screening per identificare gli obiettivi farmaco potenzialmente romanzo in Cancer Cell Culture Spheroid

The identification of functional driver events in cancer is central to furthering our understanding of cancer biology and indispensable for the discovery ...

Ricerca

Ricerca sul cancro

Invasive Behavior of Human Breast Cancer Cells in Embryonic Zebrafish

In many cases, cancer patients do not die of a primary tumor, but rather because of metastasis. Although numerous rodent models are available for studying cancer metastasis in vivo, other efficient, reliable, low-cost models are needed to quickly access the potential effects of (epi)genetic changes or pharmacological compounds. As such, we illustrate and explain the feasibility of xenograft models using human breast cancer cells injected into zebrafish embryos to support this goal. Under the microscope, fluorescent proteins or chemically labeled human breast cancer cells are transplanted into transgenic zebrafish embryos, Tg (fli:EGFP), at the perivitelline space or duct of Cuvier (Doc) 48 h after fertilization. Shortly afterwards, the temporal-spatial process of cancer cell invasion, dissemination, and metastasis in the living fish body is visualized under a fluorescent microscope. The models using different injection sites, i.e., perivitelline space or Doc are complementary to one another, reflecting the early stage (intravasation step) and late stage (extravasation step) of the multistep metastatic cascade of events. Moreover, peritumoral and intratumoral angiogenesis can be observed with the injection into the perivitelline space. The entire experimental period is no more than 8 days. These two models combine cell labeling, micro-transplantation, and fluorescence imaging techniques, enabling the rapid evaluation of cancer metastasis in response to genetic and pharmacological manipulations.

Here, we describe xenograft zebrafish models using two different injection sites, i.e., perivitelline space and duct of Cuvier, to investigate the invasive behavior and to assess the intravasation and extravasation potential of human breast cancer cells, respectively.

Invasiva comportamento delle cellule cancro al seno umano in Zebrafish embrionali

In many cases, cancer patients do not die of a primary tumor, but rather because of metastasis. Although numerous rodent models are available for studying ...

Live-imaging of Breast Epithelial Cell Migration After the Transient Depletion of TIP60

The wound-healing assay is efficient and one of the most economical ways to study cell migration in vitro. Conventionally, images are taken at the beginning and end of an experiment using a phase-contrast microscope, and the migration abilities of cells are evaluated by the closure of wounds. However, cell movement is a dynamic phenomenon, and a conventional method does not allow for tracking single-cell movement. To improve current wound-healing assays, we use live-cell imaging techniques to monitor cell migration in real time. This method allows us to determine the cell migration rate based on a cell tracking system and provides a clearer distinction between cell migration and cell proliferation. Here, we demonstrate the use of live-cell imaging in wound-healing assays to study the different migration abilities of breast epithelial cells influenced by the presence of TIP60. As cell motility is highly dynamic, our method provides more insights into the processes of wound healing than a snapshot of wound closure taken with the traditional imaging techniques used for wound-healing assays.

Here, we present the real-time monitoring of cell migration in a wound-healing assay using TIP60-depleted MCF10A breast epithelial cells. The implementation of live-cell imaging techniques in our protocol allows us to analyze and visualize single-cell movement in real time and across time.

Live-imaging della migrazione delle cellule epiteliali della mammella dopo lo svuotamento transitorio di TIP60

The wound-healing assay is efficient and one of the most economical ways to study cell migration in vitro. Conventionally, images are taken at the ...

Evaluating the Role of Mitochondrial Function in Cancer-related Fatigue

Fatigue is a common and debilitating condition that affects most cancer patients. To date, fatigue remains poorly characterized with no diagnostic test to objectively measure the severity of this condition. Here we describe an optimized method for assessing mitochondrial function of PBMCs collected from fatigued cancer patients. Using a compact extracellular flux system and sequential injection of respiratory inhibitors, we examined PBMC mitochondrial functional status by measuring basal mitochondrial respiration, spare respiratory capacity, and energy phenotype, which describes the preferred energy pathway to respond to stress. Fresh PBMCs are readily available in the clinical setting using standard phlebotomy. The entire assay described in this protocol can be completed in less than 4 hours without the involvement of complex biochemical techniques. Additionally, we describe a normalization method that is necessary for obtaining reproducible data. The simple procedure and normalization methods presented allow for repeated sample collection from the same patient and generation of reproducible data that can be compared between time points to evaluate potential treatment effects.

Our goal was to develop a practical protocol to evaluate mitochondrial dysfunction associated with fatigue in cancer patients. This innovative protocol is optimized for clinical use involving only standard phlebotomy and basic laboratory procedures.

Valutare il ruolo della funzione mitocondriale in affaticamento legato al cancro

Fatigue is a common and debilitating condition that affects most cancer patients. To date, fatigue remains poorly characterized with no diagnostic test to ...

Establishment and Characterization of Three Afatinib-resistant Lung Adenocarcinoma PC-9 Cell Lines Developed with Increasing Doses of Afatinib

Acquired resistance to molecular target inhibitors is a severe problem in cancer therapy. Lung cancer remains the leading cause of cancer-related death in most countries. The discovery of "oncogenic driver mutations," such as epidermal growth factor receptor (EGFR)-activating mutations, and subsequent development of molecular targeted agents of EGFR tyrosine kinase inhibitors (TKIs) (gefitinib, erlotinib, afatinib, dacomitinib, and osimertinib) have dramatically altered lung cancer treatment in recent decades. However, these drugs are still not effective in patients with non-small cell lung cancer (NSCLC) carrying EGFR-activating mutations. Following acquired resistance, the systemic progression of NSCLC remains a significant obstacle in treating patients with EGFR mutation-positive NSCLC. Here, we present a stepwise dose escalation method for establishing three independent acquired afatinib-resistant cell lines from NSCLC PC-9 cells harboring EGFR-activating mutations of 15-base pair deletions in EGFR exon 19. Methods for characterizing the three independent afatinib-resistance cell lines are briefly presented. The acquired resistance mechanisms to EGFR TKIs are heterogeneous. Therefore, multiple cell lines with acquired resistance to EGFR-TKIs must be examined. Ten to twelve months are required to obtain cell lines with acquired resistance using this stepwise dose escalation approach. The discovery of novel acquired resistance mechanisms will contribute to the development of more effective and safe therapeutic strategies.

A method for establishing afatinib-resistance cell lines from lung adenocarcinoma PC-9 cells was developed, and resistant cells were characterized. The resistant cells can be used to investigate epidermal growth factor receptor tyrosine kinase inhibitor-resistance mechanisms, applicable for patients with non-small cell lung cancer.

Istituzione e caratterizzazione di tre linee cellulari adenocarcinoma polmonare resistente agli Agrassiinib sviluppate con l'aumento delle dosi di Afatinib

Acquired resistance to molecular target inhibitors is a severe problem in cancer therapy. Lung cancer remains the leading cause of cancer-related death in ...

Differentiation of Mouse Breast Epithelial HC11 and EpH4 Cells

Cadherins play an important role in the regulation of cell differentiation as well as neoplasia. Here we describe the origins and methods of the induction of differentiation of two mouse breast epithelial cell lines, HC11 and EpH4, and their use to study complementary stages of mammary gland development and neoplastic transformation.
The HC11 mouse breast epithelial cell line originated from the mammary gland of a pregnant Balb/c mouse. It differentiates when grown to confluence attached to a plastic Petri dish surface in medium containing fetal calf serum and Hydrocortisone, Insulin and Prolactin (HIP medium). Under these conditions, HC11 cells produce the milk proteins &#946;-casein and whey acidic protein (WAP), similar to lactating mammary epithelial cells, and form rudimentary mammary gland-like structures termed &#34;domes&#34;.
The EpH4 cell line was derived from spontaneously immortalized mouse mammary gland epithelial cells isolated from a pregnant Balb/c mouse. Unlike HC11, EpH4 cells can fully differentiate into spheroids (also called mammospheres) when cultured under three-dimensional (3D) growth conditions in HIP medium. Cells are trypsinized, suspended in a 20% matrix consisting of a mixture of extracellular matrix proteins produced by Engelbreth-Holm-Swarm (EHS) mouse sarcoma cells, plated on top of a layer of concentrated matrix coating a plastic Petri dish or multiwell plate, and covered with a layer of 10% matrix-containing HIP medium. Under these conditions, EpH4 cells form hollow spheroids that exhibit apical-basal polarity, a hollow lumen, and produce &#946;-casein and WAP.
Using these techniques, our results demonstrated that the intensity of the cadherin/Rac signal is critical for the differentiation of HC11 cells. While Rac1 is necessary for differentiation and low levels of activated RacV12 increase differentiation, high RacV12 levels block differentiation while inducing neoplasia. In contrast, EpH4 cells represent an earlier stage in mammary epithelial differentiation, which is inhibited by even low levels of RacV12.

We describe techniques for differentiation induction of two breast epithelial lines, HC11 and EpH4. While both require fetal calf serum, insulin, and prolactin to produce milk proteins, EpH4 cells can fully differentiate into mammospheres in three-dimensional culture. These complementary models are useful for signal transduction studies of differentiation and neoplasia.

Differenziazione delle cellule epiteliali del seno del topo HC11 e EpH4

Cadherins play an important role in the regulation of cell differentiation as well as neoplasia. Here we describe the origins and methods of the induction ...

Studying Triple Negative Breast Cancer Using Orthotopic Breast Cancer Model

Triple-negative breast cancer (TNBC) is an aggressive breast cancer subtype with limited therapeutic options. When compared to patients with less aggressive breast tumors, the 5-year survival rate of TNBC patients is 77% due to their characteristic drug-resistant phenotype and metastatic burden. Toward this end, murine models have been established aimed at identifying novel therapeutic strategies limiting TNBC tumor growth and metastatic spread. This work describes a practical guide for the TNBC orthotopic model where MDA-MB-231 breast cancer cells suspended in a basement membrane matrix are implanted in the fourth mammary fat pad, which closely mimics the cancer cell behavior in humans. Measurement of tumors by caliper, lung metastasis assessment via in vivo and ex vivo imaging, and molecular detection are discussed. This model provides an excellent platform to study therapeutic efficacy and is especially suitable for the study of the interaction between the primary tumor and distal metastatic sites.

This work presets an advanced protocol to accurately assess tumor loading by detection of green fluorescent protein and bioluminescence signals as well as the integration of quantitative molecular detection technique.

Studiare il cancro al seno triplo negativo utilizzando il modello di cancro al seno ortotopico

Triple-negative breast cancer (TNBC) is an aggressive breast cancer subtype with limited therapeutic options. When compared to patients with less ...

Measuring the Motor Aspect of Cancer-Related Fatigue using a Handheld Dynamometer

Cancer-related fatigue (CRF) is commonly reported by patients both during and after receiving treatment for cancer. Current CRF diagnoses rely on self-report questionnaires which are subject to report and recall biases. Objective measurements using a handheld dynamometer, or handgrip device, have been shown in recent studies to correlate significantly with subjective self-reported fatigue scores. However, variations of both the handgrip fatigue test and fatigue index calculations exist in the literature. The lack of standardized methods limits the utilization of the handgrip fatigue test in the clinical and research settings. In this study, we provide detailed methods for administering the physical fatigue test and calculating the fatigue index. These methods should supplement existing self-reported fatigue questionnaires and help clinicians assess fatigue symptom severity in an objective and quantitative manner.

Simple and accessible methods were developed to measure the motor aspect of cancer-related fatigue objectively and quantitatively. We describe, in detail, ways to administer the physical fatigue test using a simple handgrip device as well as methods to calculate fatigue indices.

Misurazione dell'aspetto motorio della fatica correlata al cancro utilizzando un dinamometro palmo

Cancer-related fatigue (CRF) is commonly reported by patients both during and after receiving treatment for cancer. Current CRF diagnoses rely on ...

Orthotopic Transplantation of Breast Tumors as Preclinical Models for Breast Cancer

Preclinical models that faithfully recapitulate tumor heterogeneity and therapeutic response are critical for translational breast cancer research. Immortalized cell lines are easy to grow and genetically modify to study molecular mechanisms, yet the selective pressure from cell culture often leads to genetic and epigenetic alterations over time. Patient-derived xenograft (PDX) models faithfully recapitulate the heterogeneity and drug response of human breast tumors. PDX models exhibit a relatively short latency after orthotopic transplantation that facilitates the investigation of breast tumor biology and drug response. The transplantable genetically engineered mouse models allow the study of breast tumor immunity. The current protocol describes the method to orthotopically transplant breast tumor fragments into the mammary fat pad followed by drug treatments. These preclinical models provide valuable approaches to investigate breast tumor biology, drug response, biomarker discovery and mechanisms of drug resistance.

Patient-derived xenograft (PDX) models and transplantable genetically engineered mouse models faithfully recapitulate human disease and are preferred models for basic and translational breast cancer research. Here, a method is described to orthotopically transplant breast tumor fragments into the mammary fat pad to study tumor biology and evaluate drug response.

Trapianto ortotopico di tumori al seno come modelli preclinici per il cancro al seno

Preclinical models that faithfully recapitulate tumor heterogeneity and therapeutic response are critical for translational breast cancer research. ...

Modeling Breast Cancer in Human Breast Tissue using a Microphysiological System

Breast cancer (BC) remains a leading cause of death for women. Despite more than $700 million invested in BC research annually, 97% of candidate BC drugs fail clinical trials. Therefore, new models are needed to improve our understanding of the disease. The NIH Microphysiological Systems (MPS) program was developed to improve the clinical translation of basic science discoveries and promising new therapeutic strategies. Here we present a method for generating MPS for breast cancers (BC-MPS). This model adapts a previously described approach of culturing primary human white adipose tissue (WAT) by sandwiching WAT between adipose-derived stem cell sheets (ASC)s. Novel aspects of our BC-MPS include seeding BC cells into non-diseased human breast tissue (HBT) containing native extracellular matrix, mature adipocytes, resident fibroblasts, and immune cells; and sandwiching the BC-HBT admixture between HBT-derived ASC sheets. The resulting BC-MPS is stable in culture ex vivo for at least 14 days. This model system contains multiple elements of the microenvironment that influence BC including adipocytes, stromal cells, immune cells, and the extracellular matrix. Thus BC-MPS can be used to study the interactions between BC and its microenvironment. 
We demonstrate the advantages of our BC-MPS by studying two BC behaviors known to influence cancer progression and metastasis: 1) BC motility and 2) BC-HBT metabolic crosstalk. While BC motility has previously been demonstrated using intravital imaging, BC-MPS allows for high-resolution time-lapse imaging using fluorescence microscopy over several days. Furthermore, while metabolic crosstalk was previously demonstrated using BC cells and murine pre-adipocytes differentiated into immature adipocytes, our BC-MPS model is the first system to demonstrate this crosstalk between primary human mammary adipocytes and BC cells in vitro.

This protocol describes the construction of an in vitro microphysiological system for studying breast cancer using primary human breast tissue with off the shelf materials.

Modellazione del cancro al seno nel tessuto mammario umano utilizzando un sistema microfisiologico

Breast cancer (BC) remains a leading cause of death for women. Despite more than $700 million invested in BC research annually, 97% of candidate BC drugs ...