Agradecemos al Dr. Alexander Flugel (Martinsried, Alemania) para darnos la GFP-etiquetados, ovoalbúmina línea de células T específicas utilizadas en este protocolo.

1. La estimulación de los linfocitos T específicos ovoalbúmina Descanso específicos ovoalbúmina las células T son estimulados con ovoalbúmina en la presencia de ratas Lewis timocitos irradiados (30 Gy) como células presentadoras de antígeno (APC), en medio de estimulación (Preparar en DMEM + penicilina / estreptomicina / L + GLUT-AANE RPMI + Vitaminas + + 2ME suero de rata + ovoalbúmina, a las concentraciones finales se señala en la lista de materiales.) Suero de rata preparación <ol><li> Tomamos el suero de las ratas que el sacrificio como donantes timo. </li><li> Las ratas son profundamente anestesiados por inhalación de halotano y tanta sangre como sea posible es dibujado por la perfusión cardiaca (5-10 ml por rata) con jeringas de 10 ml y 23 G 1 &quot;agujas, después de que las ratas son decapitados inmediatamente para asegurar la muerte. </li><li> La sangre se deja coagular durante 10 minutos a 37 ° C seguido por 10 min en hielo. </li><li> El coágulo se extrae y se dio el resto del líquido. </li><li> El suero se recoge, filtra de manera estéril y almacenadas a -20 ° C. </li></ol> Preparación de las células presentadoras de antígeno <ol><li> Tome el timo de una rata decapitado con instrumentos estériles y colocarlo en PBS con penicilina y estreptomicina (PBS-PS), sobre el hielo. </li><li> Tome el timo a una campana de cultivo de tejidos, lo limpia de coágulos sanguíneos y otros tejidos contaminantes y cortar en trozos pequeños en un colador de la célula (Fisher # 08-771-2) colocada en una placa de Petri de 10 cm con 10 ml de PBS- PS. </li><li> Cada pieza se presiona a través del filtro de células usando la parte de atrás de un estéril de 1 ml émbolo de la jeringa. Recoger la única suspensión de células en un tubo de 50 ml en hielo. </li><li> Enjuague el filtro celular con 20-40 ml más de PBS-PS. Centrifugar la suspensión a 1250g y resuspender el precipitado en PBS-PS, 5 ml / timo, en el hielo. </li><li> Irradiar esta suspensión (30 Gy) en un irradiador gamma e inmediatamente lavar dos veces con 50 ml de PBS-PS. Nota: Los mejores donantes del timo son las ratas jóvenes adultos, de 5-7 semanas de edad. Las ratas de hasta 10 semanas puede ser utilizado, pero el número de células disminuye con el tiempo. El género no es importante, aunque algunas personas dicen que los hombres son mejores donantes del timo. Siempre usamos las mujeres, porque todos nuestros animales están alojados en la misma habitación y no queremos que hombres y mujeres en la misma habitación. </li></ol> Estímulo <ol><li> Lave los linfocitos T una vez y contar con ellos. </li><li> Contar las células presentadoras de antígeno. </li><li> Mezcle 3 x 10 6 ovoalbúmina específicos de las células T de 150 x 10 6 vehículos blindados en una placa de Petri de 10 cm con 10 ml de medio de estimulación. </li><li> Incubar durante 48 horas a 37 ° C. </li></ol> 2. La transferencia adoptiva de los linfocitos T específicos ovoalbúmina <ol><li> Las ratas que utilizamos como receptores son mujeres 70-10 semanas de edad. </li><li> Contar con los linfocitos T, que debe ser grande y redondo. Las células presentadoras de antígeno estará muerto. </li><li> Centrifugar los linfocitos T y volver a suspender las de 10 x 10 6 células / ml en PBS. </li><li> Inyectar 1 ml por vía intraperitoneal con jeringas de 3 ml y 23 G 1 agujas &quot;. </li></ol> 3. Desafío con el antígeno en el oído <ol><li> Dos días después de la transferencia adoptiva, se inyectan 20 l de una solución de 1 mg / ml ovoalbúmina (= antígeno relevante) por un oído y solución salina (o antígeno irrelevante) en el otro oído. No filtrar la solución de antígeno, ya que puede perder los agregados que potencian la reacción inflamatoria. Nota: Las ratas deben ser anestesiados para evitar movimientos. Si usted es diestro, mantenga la oreja entre el dedo pulgar izquierdo y el dedo índice, el dedo índice está dentro del oído y de la rata frente a usted. </li><li> Se inyecta la solución en el pabellón auricular de la oreja utilizando 27G ½ &quot;agujas. Trate de inyectarse a todas las ratas exactamente en el mismo lugar. </li></ol> 4. La medición de DTH <ol><li> El oído comenzará a girar roja e hinchada como un signo de inflamación. Medir el grosor de cada oído con un micrómetro de resorte (por ejemplo modelo PK-0505 de Mitutoyo). </li><li> Tome 5-6 mediciones para cada oído en cada punto del tiempo y calcular la media. </li><li> La inflamación máximo debe ocurrir entre 24 y 72 horas. </li></ol>

<ol>
	<li>Beeton, C., Barbaria, J., Devaux, J., Benoliel, A. -. M., Gola, M., Sabatier, J. -. M., Bernard, D., Crest, M., Beraud, E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Selective+blocking+of+voltage-gated+K++channels+treats+experimental+autoimmune+encephalomyelitis+and+inhibits+T-cell+activation.">Selective blocking of voltage-gated K+ channels treats experimental autoimmune encephalomyelitis and inhibits T-cell activation.</a> J. Immunol. , 166-936 (2001).</li><li>Beeton, C., Chandy, K. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Induction+and+monitoring+of+active+delayed+type+hypersensitivity+(DTH)in+rats.">Induction and monitoring of active delayed type hypersensitivity (DTH)in rats.</a> J Vis Exp. (5), (2007).</li></ol>

El desafío antigénico también se puede realizar en otros sitios, por ejemplo en la piel de la espalda (2). Sin embargo nos encontramos que un reto. Usando el pabellón de la oreja, como se muestra en este video, permite la medición más precisa de la reacción de DTH.

<table><tbody><tr><td>DMEM</td><td>GIBCO, by Life Technologies</td><td>12430-062</td><td>500ml</td></tr><tr><td>Sodium pyruvate</td><td>Sigma-Aldrich</td><td>S8636</td><td>1 mM final, in Stimulation medium</td></tr><tr><td>Penicillin &amp;#8211; Streptomycin &amp;#8211; L-Glutamine</td><td>Cambrex/BioWhittaker</td><td>17-718R</td><td>100 u/ml - 100 &amp;#x03BC;g/ml - 4 mM respectively, final, in Stimulation Medium </td></tr><tr><td>Non essential amino acids</td><td>Sigma-Aldrich</td><td>R7131</td><td>NEAA, 1% final, in Stimulation Medium</td></tr><tr><td>RPMI vitamins</td><td>Sigma-Aldrich</td><td>R7256</td><td>1% final, in Stimulation medium</td></tr><tr><td>beta-mercapt&amp;#8211;thanol</td><td>Sigma-Aldrich</td><td>M6250</td><td>2ME, 50 &amp;#x03BC;M final, in Stimulation Medium</td></tr><tr><td>Syngeneic rat serum</td><td></td><td></td><td>1% final. See explanation in Protocol.</td></tr><tr><td>Ovalbumin</td><td>Sigma-Aldrich</td><td>A5503</td><td>10 &amp;#x03BC;g/ml final, in Stimulation Medium.</td></tr></tbody></table>

induction and monitoring of adoptive delayed-type hypersensitivity in rats

Retardada de hipersensibilidad (DTH) es una reacción inflamatoria mediada por CCR7-efector linfocitos T de memoria que se infiltran en el sitio de la inyección de un antígeno contra el que ha sido el sistema inmunológico preparado. La reacción inflamatoria se caracteriza por enrojecimiento e inflamación del sitio de estímulos antigénicos. Es un modelo práctico para determinar la eficacia in vivo de los inmunosupresores. DTH cutánea puede ser inducido, ya sea por transferencia adoptiva de linfocitos T antígeno-específicos o mediante la inmunización activa con un antígeno, y el desafío posterior intradérmica con el antígeno para inducir la reacción inflamatoria en la piel de una zona determinada. Respuestas DTH pueden ser inducidas a diversos antígenos, para la ovoalbúmina ejemplo, la tuberculina, toxoide tetánico, o hemocianina de lapa californiana. Estas reacciones también pueden ser inducidos en contra de autoantígenos, por ejemplo, proteína básica de mielina (MBP) en ratas con encefalomielitis autoinmune experimental inducida con MBP, un modelo animal de esclerosis múltiple (1). Aquí se demuestra la forma de inducir una reacción adoptivos DTH en ratas Lewis. En primer lugar, se estimula la ovoalbúmina específicos de las células T in vitro y se inyecta por vía intraperitoneal estas células activadas a las ratas ingenuo. Después de permitir que las células que se equilibre en vivo durante 2 días, le desafío a las ratas con ovoalbúmina en el pabellón de una oreja, mientras que el otro oído Wil reciben solución salina. La reacción inflamatoria será visible 3-72 horas más tarde y espesor de la oreja se medirá como una indicación de la gravedad de DTH.

Watch this Scientific Journal Video about Induction and Monitoring of Adoptive Delayed-Type Hypersensitivity in Rats at JoVE.com

Induction and Monitoring of Adoptive Delayed-Type Hypersensitivity in Rats

Retardada de hipersensibilidad (DTH) es una reacción inflamatoria mediada por CCR7-T de memoria efectora (TEM) de linfocitos. Aquí se demuestra cómo activar antígenos específicos de células TEM, inducir DTH adoptivos en ratas Lewis y controlar la respuesta inflamatoria.

Inducción y Seguimiento de Adopción de hipersensibilidad retardada en ratas

Delayed type hypersensitivity (DTH) is an inflammatory reaction mediated by CCR7- effector memory T lymphocytes that infiltrate the site of injection of ...

induction-monitoring-adoptive-delayed-type-hypersensitivity

Research

JoVE Journal

Biology

11.9K vistas.  University of California, Irvine (UCI). Retardada de hipersensibilidad (DTH) es una reacción inflamatoria mediada por CCR7-T de memoria efectora (TEM) de linfocitos. Aquí se demuestra cómo activar antígenos específicos de células TEM, inducir DTH adoptivos en ratas Lewis y controlar la respuesta inflamatoria.

Video: Induction and Monitoring of Adoptive Delayed-Type Hypersensitivity in Rats

Induction and Clinical Scoring of Chronic-Relapsing Experimental Autoimmune Encephalomyelitis

Multiple sclerosis (MS) is a chronic inflammatory disease of the central nervous system (CNS) that commonly affects young adults. It is characterized by demyelination and glial scaring in areas disseminated in the brain and spinal cord. These lesions alter nerve conduction and induce the disabling neurological deficits that vary with the location of the demyelinated plaques in the CNS (e.g. paraparesis, paralysis, blindness, incontinence).
Experimental autoimmune encephalomyelitis (EAE) is a model for MS. EAE was first induced accidentally in humans during vaccination against rabies, using viruses grown on rabbit spinal cords. Residues of spinal injected with the inactivated virus induced the CNS disease. Following these observations, a first model of EAE was described in non-human primates immunized with a CNS homogenate by Rivers and Schwenther in 1935. EAE has since been generated in a variety of species and can follow different courses depending on the species/strain and immunizing antigen used. For example, immunizing Lewis rats with myelin basic protein in emulsion with adjuvant induces an acute model of EAE, while the same antigen induces a chronic disease in guinea pigs.
The EAE model described here is induced by immunizing DA rats against DA rat spinal cord in emulsion in complete Freund's adjuvant. Rats develop an ascending flaccid paralysis within 7-14 days post-immunization. Clinical signs follow a relapsing-remitting course over several weeks. Pathology shows large immune infiltrates in the CNS and demyelination plaques. Special considerations for taking care for animals with EAE are described at the end of the video.

This video demonstrates the induction and clinical scoring of an animal model of multiple sclerosis: chronic-relapsing experimental autoimmune encephalomyelitis in DA rats. The disease, induced by immunizing rats with an emulsion containing whole rat spinal cord and complete Freund's adjuvant, presents clinical signs resembling the human disease.

La inducción y la puntuación clínica de enfermedades crónico-recurrente encefalomielitis autoinmune experimental

Multiple sclerosis (MS) is a chronic inflammatory disease of the central nervous system (CNS) that commonly affects young adults. It is characterized by ...

Investigación

Biología

Induction and Monitoring of Active Delayed Type Hypersensitivity (DTH) in Rats

Delayed type hypersensitivity (DTH) is an inflammatory reaction mediated by CCR7- effector memory T lymphocytes that infiltrate the site of injection of an antigen against which the immune system has been primed. The inflammatory reaction is characterized by redness and swelling of the site of antigenic challenge. It is a convenient model to determine the in vivo efficacy of immunosuppressants. Cutaneous DTH can be induced either by adoptive transfer of antigen-specific T lymphocytes or by active immunization with an antigen, and subsequent intradermal challenge with the antigen to induce the inflammatory reaction in a given skin area. DTH responses can be induced to various antigens, for example ovalbumin, tuberculin, tetanus toxoid, or keyhole limpet hemocyanin (KLH).
Here we demonstrate how to induce an active DTH reaction in Lewis rats. We will first prepare a water-in-oil emulsion of KLH, our antigen of interest, in complete Freund's adjuvant and inject this emulsion subcutaneously to rats. This will prime the immune system to develop memory T cells directed to KLH. Seven days later we will challenge the rats intradermally on the back with KLH on one side and with ovalbumin, an irrelevant antigen, on the other side. The inflammatory reaction will be visible 16-72 hours later and the red and swollen area will be measured as an indication of DTH severity.

Induction and Monitoring of Active Delayed Type Hypersensitivity DTH in Rats

Delayed type hypersensitivity (DTH) is an inflammatory reaction mediated by CCR7- effector memory T lymphocytes. Here we demonstrate how to induce active DTH in Lewis rats and monitor the inflammatory response.

Inducción y seguimiento de activos de hipersensibilidad de tipo retardado (DTH) en ratas

Drawing Blood from Rats through the Saphenous Vein and by Cardiac Puncture

Drawing blood from rodents is necessary for a large number of both in vitro and in vivo studies. Sites of blood draws are numerous in rodents: retro-orbital sinus, jugular vein, maxillary vein, saphenous vein, heart. Each technique has its advantages and disadvantages, and some are not approved any more in some countries (e.g., retro-orbital draws in Holland). A discussion of different techniques for drawing blood are available 1-3.
Here, we present two techniques for drawing blood from rats, each with its specific applications.
Blood draw from the saphenous vein, provided it is done properly, induces minimal distress in animals and does not require anesthesia. This technique allows repeated draws of small amounts of blood, such as needed for pharmacokinetic studies 4,5, determining plasma chemistry, or blood counts 6.
Cardiac puncture allows the collection of large amounts of blood from a single animal (up to 10 ml of blood can be drawn from a 150 g rat). This technique is therefore very useful as a terminal procedure when drawing blood from the saphenous would not provide a large enough sample. We use cardiac puncture when we need sufficient amounts of serum from a specific strain of rats to grow T lymphocyte lines in vitro 4-9.

Blood draws are necessary in a large number of studies, for example to study the pharmacokinetics profile of a compound. Here, we demonstrate how to draw blood from rats using two techniques: blood draw from the saphenous vein or by cardiac puncture.

La extracción de sangre de las ratas a través de la vena safena y por punción cardiaca

Drawing blood from rodents is necessary for a large number of both in vitro and in vivo studies. Sites of blood draws are numerous in rodents: ...

Imaging Effector Memory T cells in the Ear After Induction of Adoptive DTH

Delayed type hypersensitivity (DTH) is an immune reaction in which the main players are CCR7- effector / memory T lymphocytes. Here, we demonstrate a method for inducing and recording the progress of a DTH reaction in the rat ear. This is followed by a demonstration of the preparation of rat ear tissue for two-photon imaging of the CCR7- effector / memory T cell response.
An adoptive DTH is induced by the intraperitoneal injection of GFP-labeled Ova-specific CCR7- effector / memory T cell line (Beeton, C J. Visualized Experiments, Issue 8). Cells are then allowed to equilibrate in the rat for 48 hours before challenge by injecting one ear with saline (control ear) and the other with a 1:1 mix of Ova and Ova conjugated to Texas-Red (Ova-TR) to allow visualization of resident antigen-presenting cells.
We describe a method of tissue preparation useful for imaging the motility of cells within the deep dermal layer during an immune response, in conjunction with visualization of collagen fibers by second harmonic generation. Ear tissue is cut into 5 x 5 mm squares (slightly larger is better) and mounted onto plastic cover slips using Vetbond&trade;, which are then secured using silicone grease in an imaging chamber and superfused by oxygen-bubbled tissue culture medium at 37&deg;C.

Here we demonstrate a method for inducing and recording the progress of a delayed type-hypersensitivity (DTH) reaction in the rat ear. This is followed by a demonstration of the preparation of rat ear tissue for two-photon imaging of the effector / memory T cell response.

Imágenes de las células efectoras T de memoria en el oído después de la inducción de DTH adoptivos

Delayed type hypersensitivity (DTH) is an immune reaction in which the main players are CCR7- effector / memory T lymphocytes. Here, we demonstrate a ...

Trans-vivo Delayed Type Hypersensitivity Assay for Antigen Specific Regulation

Delayed-type hypersensitivity response (DTH) is a rapid in vivo manifestation of T cell-dependent immune response to a foreign antigen (Ag) that the host immune system has experienced in the recent past. DTH reactions are often divided into a sensitization phase, referring to the initial antigen experience, and a challenge phase, which usually follows several days after sensitization. The lack of a delayed-type hypersensitivity response to a recall Ag demonstrated by skin testing is often regarded as an evidence of anergy. The traditional DTH assay has been effectively used in diagnosing many microbial infections.
Despite sharing similar immune features such as lymphocyte infiltration, edema, and tissue necrosis, the direct DTH is not a feasible diagnostic technique in transplant patients because of the possibility of direct injection resulting in sensitization to donor antigens and graft loss. To avoid this problem, the human-to-mouse "trans-vivo" DTH assay was developed 1,2. This test is essentially a transfer DTH assay, in which human peripheral blood mononuclear cells (PBMCs) and specific antigens were injected subcutaneously into the pinnae or footpad of a na&iuml;ve mouse and DTH-like swelling is measured after 18-24 hr 3. The antigen presentation by human antigen presenting cells such as macrophages or DCs to T cells in highly vascular mouse tissue triggers the inflammatory cascade and attracts mouse immune cells resulting in swelling responses. The response is antigen-specific and requires prior antigen sensitization. A positive donor-reactive DTH response in the Tv-DTH assay reflects that the transplant patient has developed a pro-inflammatory immune disposition toward graft alloantigens.
The most important feature of this assay is that it can also be used to detect regulatory T cells, which cause bystander suppression. Bystander suppression of a DTH recall response in the presence of donor antigen is characteristic of transplant recipients with accepted allografts 2,4-14. The monitoring of transplant recipients for alloreactivity and regulation by Tv-DTH may identify a subset of patients who could benefit from reduction of immunosuppression without elevated risk of rejection or deteriorating renal function.
A promising area is the application of the Tv-DTH assay in monitoring of autoimmunity15,16 and also in tumor immunology 17.

We describe a valuable diagnostic assay that could potentially be used to decide the withdrawal of immunosuppression after transplant without elevated risk of graft rejection. The assay uses the principles of Delayed Type Hypersensitivity and provides accurate assessment of both donor specific effector and regulatory immune responses mounted by recipients.

Trans-vivo hipersensibilidad de tipo retardada Ensayo para Antigen Reglamento Específico

Delayed-type hypersensitivity response (DTH) is a rapid in vivo manifestation of T cell-dependent immune response to a foreign antigen (Ag) that the host ...

Inmunología e Infección

Effects of Exposure of Formaldehyde to a Rat Model of Atopic Dermatitis Induced by Neonatal Capsaicin Treatment

Atopic dermatitis is chronically relapsing pruritic eczema and prevails around the world especially in developed countries. Complex interactions between genetic and environmental factors are known to play an important role in the pathophysiology of atopic dermatitis. However, we still lack a detailed picture of the pathogenesis of this disease. Thus, it is of importance to develop appropriate animal models for elucidating the progression of atopic dermatitis. Moreover, investigating the effect of environmental factors such as air pollutants on atopic dermatitis expands understanding of the disease. Here, we describe a method for inducing atopic dermatitis in rats with neonatal capsaicin treatment and a protocol for exposure of a constant concentration of formaldehyde to rats to reveal effects on the development of atopic dermatitis in infantile and adolescent periods. These protocols have been successfully applied to several experiments and can be used for other substances.

We describe here methods for neonatal capsaicin treatment to induce atopic dermatitis in rats and exposure of vaporized formaldehyde to investigate the effects of formaldehyde inhalation on atopic dermatitis.

Efectos de la exposición de formaldehído a un modelo de rata de la Dermatitis atópica inducida por el tratamiento Neonatal de capsaicina

Atopic dermatitis is chronically relapsing pruritic eczema and prevails around the world especially in developed countries. Complex interactions between ...

Mouse Body Temperature Measurement Using Infrared Thermometer During Passive Systemic Anaphylaxis and Food Allergy Evaluation

Mouse body temperature measurement is of paramount importance for investigating allergies and anaphylactic symptoms. Rectal probes for temperature readings is common, and they have been proven to be accurate and invaluable in this regard. However, this method of temperature measurement requires the mice to be anesthetized in order to insert the probe without injury to the animal. This limits the ability to observe other phenotypes of the mouse simultaneously. In order to investigate other phenotypes while measuring temperatures, rectal probes are not ideal, and another method is desired. Here, we introduce a noninvasive method of temperature measurement that foregoes the requirement for mouse anesthesia while maintaining equal reliability to rectal probes in measuring body temperature. We use an infrared thermometer that detects body surface temperatures at ranges between 2 and 150 mm. This method of body temperature measurement is successful in reliably replicating temperature change trends during passive system anaphylaxis experiments in mice. We show that body surface temperatures are about 2.0 &#176;C lower than rectal probe measurements, but the degree of temperature drop follows the same trend. Furthermore, we use the same technique to observe mice in a food allergy model to evaluate temperature and activity levels simultaneously.

Here we present a new method to accurately measure body temperature differences in passive systemic anaphylaxis (PSA) and food allergy mouse models using an infrared thermometer. This procedure has been accurately duplicated in previous PSA results.

Medición de la temperatura del cuerpo de ratón con termómetro infrarrojo pasiva Anafilaxia sistémica y evaluación de la alergia alimentaria

Mouse body temperature measurement is of paramount importance for investigating allergies and anaphylactic symptoms. Rectal probes for temperature ...

Induction of Drug-Induced, Autoimmune Hepatitis in BALB/c Mice for the Study of Its Pathogenic Mechanisms

Drug-induced autoimmune hepatitis (DIH) is the most common hepatic drug-induced hypersensitization process observed in approximately 9 to 12% of patients with autoimmune hepatitis. The overwhelming majority of patients with DIH are women. The underlying mechanisms of these sex differences in prevalence are unclear because of the paucity of animal models that mimic human disease. Even so, underlying mechanisms are widely believed to be associated with human leukocyte antigen haplotypes and sex hormones. In contrast, using a DIH mouse model, we have uncovered that IL-4 initiated&#160;CD4+ T cells directed against an epitope of cytochrome P450 2E1 induces influx of neutrophils, macrophages and mast cells into the livers of female BALB/c mice. Using this model, we have also shown that IL-33-induced FoxP3+regulatory T cells confer protection against DIH in female and male mice. This DIH model is induced by immunizing mice with an epitope of CYP2E1 that has been covalently altered with a drug metabolite that has been associated with DIH. This epitope is recognized by patients with DIH. Our method induces robust and reproducible hepatitis and autoantibodies that can be utilized to study the pathogenesis of DIH. While in vivo studies can cause undue pain and distress in mice when done improperly, the advantage of an in vivo model is the ability to evaluate the pathogenesis of disease in a large number of mice. Additionally, biological effects of the altered liver proteins can be studied using invasive procedures. The addition of in vitro studies to the experimental design allows rapid repetition and mechanistic analysis at a cellular level. Thus, we will demonstrate our model protocol and how it can be utilized to study in vivo and in vitro mechanisms of DIH.

We describe an in vivo immunization, translational hepatitis model in BALB/c mice that can be utilized to study the pathogenesis of drug-induced autoimmune hepatitis including sex differences seen in this disease. We will describe how this model demonstrates reproducible analyses using in vivo and in vitro experimental techniques.

Inducción de hepatitis autoinmune inducida por fármacos en ratones BALB/c para el estudio de sus mecanismos patógenos

Drug-induced autoimmune hepatitis (DIH) is the most common hepatic drug-induced hypersensitization process observed in approximately 9 to 12% of patients ...

Contact Hypersensitivity as a Murine Model of Allergic Contact Dermatitis

Contact hypersensitivity (CHS) is an experimental model of allergic contact dermatitis (ACD) that can be studied in mice. This study aims to present an objective laboratory method that may help to study the CHS reaction in mice, which can be measured and quantified by various tests. To induce CHS, on day "0", mice were sensitized on a previously shaved spot by abdominal skin painting with the hapten 2,4,6-trinitrochlorobenzene (TNCB) in an acetone-ethanol mixture, whereas negative control mice were sham sensitized with vehicle alone-acetone-ethanol mixture. On day "4", the baseline ear thickness was measured with a micrometer prior to the elicitation of CHS (challenge) by painting both ears with diluted TNCB both in the test and control groups. After 24 h, the ear swelling was measured with a micrometer. CHS is an example of a T cell-mediated immune response that causes swelling in inflamed tissue, peaking 24 h after the skin challenge with the same hapten. An increase in ear edema correlated with augmented ear weight, myeloperoxidase (MPO) activity, pro-inflammatory cytokine concentration in the ear extracts, increased thickening of the edematous dermis in the histological examination, and ear vascular permeability. There was also an increase in the concentration of TNP-specific IgG1 antibodies in the sera of the test group when compared with the control mice. Additionally, CHS can be successfully transferred with the CHS-effector cells obtained from donors previously sensitized with TNCB. The CHS-effector cells were administered intravenously into na&#239;ve recipient mice, which were subsequently challenged with the same diluted hapten. Ear swelling was measured with a micrometer 24 h later.

Contact hypersensitivity (CHS) is a murine experimental model of allergic contact dermatitis (ACD). CHS is based on sensitization with reactive hapten by painting the shaved skin of the chest and abdomen, with a subsequent ear skin challenge with a diluted hapten, causing a swelling reaction that is assessed in various ways.

Hipersensibilidad de contacto como modelo murino de dermatitis alérgica de contacto

Contact hypersensitivity (CHS) is an experimental model of allergic contact dermatitis (ACD) that can be studied in mice. This study aims to present an ...

<meta charset="utf8"></head><body>Creación de un modelo de rata prematura para la investigación del dolor posparto temprano

A Preterm Rat Model for Pain Studies

This research delves into the consequences of consistent pinprick stimulation on preterm offspring to ascertain its long-term implications for pain sensitivity. The primary objective of this protocol was to investigate the impact of neonatal pinprick stimuli on the pain threshold in the later stages of life using a preterm rat model. By establishing this model, we aim to advance the research on understanding and managing early postnatal pain associated with prematurity. The findings of this study indicate that while the baseline thresholds to mechanical stimuli remained unaffected, there was a notable increase in mechanical hypersensitivity following complete Freund's adjuvant (CFA) injection in adult rats. Interestingly, compared with male rats, female rats demonstrated heightened inflammatory hypersensitivity. Notably, maternal behavior, the weight of the litters, and the growth trajectory of the offspring remained unchanged by the stimulation. The manifestation of altered nociceptive responses in adulthood after neonatal painful stimuli could be indicative of changes in sensory processing and the functioning of glucocorticoid receptors. However, further research is needed to understand the underlying mechanisms involved and to develop interventions for the consequences of prematurity and neonatal pain in adults.

<meta charset="utf8"></head><body>Un modelo de rata prematura para estudios del dolor

Here, we present a concise protocol for creating a preterm rat model, facilitating research on early postnatal pain management. The method involves performing a cesarean section three days before the expected birth, extracting preterm rat pups via hysterectomy, and integrating them with the biological offspring of a surrogate dam.

Un modelo de rata prematura para estudios del dolor

This research delves into the consequences of consistent pinprick stimulation on preterm offspring to ascertain its long-term implications for pain ...