Risk factors for complications after emergency surgery for paediatric appendicitis: a national prospective observational cohort study.

Appendicectomy is a common procedure in children with a low risk of mortality, however, complication rates and risk factors are largely unknown. This study aimed to characterise the incidence and epidemiology of postoperative complications in children undergoing appendicectomy in the UK. This multicentre prospective observational cohort study, which included children aged 1-16 y who underwent surgery for suspected appendicitis, was conducted between November 2019 and January 2022. The primary outcome was 30-day postoperative morbidity. Data collected included: patient characteristics; comorbidities; and physiological status. Multivariable regression analysis was used to identify independent risk factors for poor outcomes. Data from 2799 children recruited from 80 hospitals were analysed, of which 185 (7%) developed postoperative complications. Children from black and 'other' minority ethnic groups were at significantly higher risk of poor outcomes: OR (95%CI) 4.13 (1.87-9.08), p < 0.001 and 2.08 (1.12-3.87), p = 0.021, respectively. This finding was independent of socio-economic status and type of appendicitis found on histology. Other risk factors for complications included: ASA physical status ≥ 3 (OR (95%CI) 4.05 (1.70-9.67), p = 0.002); raised C-reactive protein (OR 95%CI 1.01 (1.00-1.01), p < 0.001); pyrexia (OR (95%CI) 1.77(1.20-2.63), p = 0.004); and peri-operative oxygen supplementation (OR (95%CI) 4.20 (1.44-12.24), p = 0.009). In the UK NHS, which is a universally accessible healthcare system, ethnicity, but not socio-economic status, was associated with an increased risk of postoperative complications in children having surgery for acute appendicitis. Further evaluations and interventions are required to address this health inequality in keeping with NHS and international priorities.

Biological Activity of Mouse OX40L-Igg Purified With Two Novel Resins.

One of the important stimulating molecules for the function of T lymphocytes is tumor necrosis factor receptor OX40 (CD134), activated by its cognate ligand OX40L (CD134L, CD252). OX40L interactions have been proposed as a potential therapeutic target for treating infectious and non-infectious diseases. The main purpose of this study was to determine the potency of two novel resins MBI and MEP for the purification of OX40L-IgG fusion protein and the biological activities of this OX40L-IgG fusion protein. The biological activity of the OX40L-IgG purified by these resins compared with protein A sepharose resin. Mice treated with the same doses of the OX40L purified by the three resins showed a significant delay in tumor growth compared to the controls injected with PBS. Mice treated with the OX40L purified by MBI resin showed a significant delay in tumor cell (CT26) growth compared with mice injected with OX40L purified by other resins.

Acretismo placentario: un diagnóstico emergente. Abordaje quirúrgico no conservador / Placental accreta: an emerging diagnosis. Approach non-conservative surgical

RESUMEN El espectro de acretismo placentario es un fenómeno infrecuente del embarazo cuya incidencia ha aumentado considerablemente y que está caracterizado por el anclaje anormal de las vellosidades coriónicas al miometrio, lo cual aumenta la morbi-mortalidad materna durante la resolución quirúrgica. Según las capas uterinas comprometidas, serán clasificadas como placenta acreta (contacta miometrio), increta (penetra miometrio) y percreta (compromete todo el miometrio y/o eventualmente órganos adyacentes), siendo su mayor factor de riesgo: la cesárea anterior y la placenta previa. En este artículo se realizó una revisión bibliográfica abarcando definiciones, diagnóstico y las nuevas tendencias en manejo quirúrgico no conservador propuesto en la nueva guía de la Federación Internacional de Ginecología y Obstetricia publicada en 2018 y elaborando una discusión respecto a ellas.

SUMMARY Placenta accreta spectrum is an uncommon phenomenon of pregnancy whose incidence has increased considerably over time and is characterized by the abnormal anchoring of the chorionic villi to the myometrium, which increases maternal morbidity and mortality during surgical resolution. According to the compromised uterine layers, they will be classified as placenta accreta (contacts myometrium), increta (penetrates myometrium) and percreta (compromises the entire myometrium and / or possibly adjacent organs), being previous caesarean section and placenta previous its major risk factor. In this review, we included definitions, diagnosis, and the new topics in non-conservative surgical management developed by the International Federation of Obstetrics and Gynecolgy published in 2018, and developing a discussion of the topic.

Utilidad de la detección de virus papiloma de alto riesgo en pacientes con citología atípica y de neoplasia intraepitelial de bajo grado de cuello uterino

Antecedentes: La citología cervical atípica y de bajo grado son hallazgos frecuentes pero no siempre relevantes. Objetivo: Estudio prospectivo que analiza la utilidad de la determinación del virus papiloma humano (VPH) de alto riesgo, en detectar patología intraepitelial de alto grado. Método: 94 pacientes con citología atípica y 116 pacientes con citología de NIE 1 fueron evaluadas con la prueba de VPH de alto riesgo. Resultados: En pacientes con citología atípica, la prueba positiva de VPH identificó a 23% con patología de alto grado versus solo en 4% en aquellas con prueba negativa (p=0,0124). En pacientes con citología de NIE 1, la prueba positiva de VPH identificó a 13% con patología de alto grado versus en 5,5% en aquellas con prueba negativa (p=0,6923). Conclusiones: La determinación de presencia viral de alto riesgo es un examen útil en pacientes con citología atípica pero no contribuye en pacientes con citología de NIE 1.

Background: Atypical and low grade cervical cytology are frequent findings, however, not always relevant. Aims: Prospective study, that analyzes the usefulness of determining high risk human papilloma virus test (HPV), in detecting high grade intra epithelial neoplasia in these patients. Method: 94 patients with atypical cytology and 116 patients with CIN 1 cytology were evaluated with the high risk HPV test. Results: In patients with atypical cytology, a positive HPV test identified 23% with high grade CIN versus only in 4% when the test was negative (p=0.0124). In patients with CIN 1 cytology, a positive test identified 13% with high grade CIN versus 5.5% in those with a negative test (p=0.6923). Conclusion: High risk HPV determination is a useful test in patients with atypical cytology, however non contributory in patients with low grade CIN cytology.

Caesarean hysterectomy for placenta praevia/accreta using an approach via the pouch of Douglas.

UNLABELLED: Placenta praevia/accreta is associated with significant maternal morbidity and mortality and is a common cause of obstetric hysterectomy. This paper describes posterior retrograde abdominal hysterectomy, a new surgical technique for caesarean hysterectomy, in 11 women with placenta percreta, increta or accreta There were no intraoperative or postoperative maternal complications, and only one fetus required admission to the neonatal unit, for prematurity. Our technique in placenta praevia/accreta allows easy identification of the vagina and early uterine devascularisation, as well as safe resection of the involved urinary bladder in women with placenta percreta showing bladder penetration. Analytical studies are needed to confirm our findings. TWEETABLE ABSTRACT: Posterior retrograde abdominal hysterectomy in women with placenta praevia/accreta may enable safer surgery.

CTL responses to DCs stimulated with leishmania antigens detected by DCs expressing Leishmania gp63.

BACKGROUND: Leishmania is a pathogenic parasite which infects mononuclear cells in vertebrate hosts. Different strategies have been taken to develop immunity against Leishmania. DCs loaded with immunogenic antigen have resulted in different levels of Th1-type immune response and cytotoxic T lymphocytes (CTL) activity. OBJECTIVE: To evaluate the potency of DCs primed with soluble Leishmania mexicana antigens (SLA) in developing CTL activity. METHODS: DCs were loaded with SLA and injected to Balb/c mice. After two weeks the mice were sacrificed and their splenocytes were used as effector cells in a standard 4-hour cytotoxicity assay against DCs transfected with pcDNA3 containing L. mexicana gp63 gene. RESULTS: Immunization of Balb/c mice with DCs loaded with SLA resulted in high levels of CTL activity against DCs transfected with pcDNA3 containing L. mexicana gp63 gene. CONCLUSIONS: The results indicate a high potency for DCs primed with Leishmania antigens in inducing CTL activity, which can be used for developing an immunogenic vaccine against Leishmania.

Uso de bloqueo paravertebral torácico bilateral en cirugía cardiaca para extubación postoperatoria inmediata: serie de casos clínicos / Use of paravertebral block thoracic bilateral in cardiac surgery to extubation Immediate postoperative: clinical case series

El desarrollo de protocolos de extubación precoz en cirugía cardiaca se basa en el uso de técnicas anestésicas que persiguen el objetivo de tener un paciente con un estado de vigilia adecuado y una vía aérea segura durante el período de emergencia de la anestesia. Se diseñó un estudio prospectivo y descriptivo con el objetivo de evaluar si el uso de bloqueo paravertebral torácico bilateral (BPVTB) previo a una cirugía cardiaca en adultos disminuye los requerimientos intraoperatorios (IO) de opioides, y aumentando con ello la posibilidad de extubación segura en el pabellón y, además, evaluar la calidad analgésica postoperatoria (PO). Material y Método: Los pacientes deben ser adultos sometidos a una cirugía cardiaca por esternotomía. Los criterios de exclusión fueron: cirugía larga y compleja, paro circulatorio, hipotermia profunda, insuficiencia cardiaca, hipertensión pulmonar severa, obesidad > grado II, enfermedad pulmonar severa y coagulopatía. Los pacientes seleccionados fueron sometidos a un BPVTB, y luego se indujo anestesia general no estandarizada. Tras realizar la cirugía prevista, al inicio del cierre de la piel se suspende el agente anestésico inhalatorio, se realiza prueba de ventilación espontánea y se extuba en pabellón si cumple criterios establecidos. Posteriormente se evaluó PaO2 y PaCO2(ingreso UCI), se aplicó escala visual análoga (EVA) para medir intensidad del dolor (6 h del PO) y al finalizar el primer día de PO se consignó el consumo de opioides, la incidencia de náuseas y vómitos y la necesidad de usar ventilación mecánica invasiva (VMI) o no invasiva (VMNI)...

The development of early extubation protocols in cardiac surgery are based on anesthesic techniques able to have the appropriate anesthesia depth and a secured airway during the emergence of anesthesia. A prospective study was design in adults subjected to cardiac surgery in order to evaluate the effectiveness of preoperative bilateral paravertebral thoracic block (BPVTB) in decreasing intraoperative opiods doses and allow a safe extubation in the operating room (OR) and, even more, give a good quality postoperative (PO) analgesia. Material and Methods: Patients subjected to a cardiac surgery with sternotomy. Exclusion criteria were: long and complex surgery, circulatory arrest, deep hypothermia, cardiac failure, severe pulmonary hypertension, obesity grade II or greater, severe lung disease and coagulophaty. A cohort of patients were subjected to a BPVTB, then general anesthesia was induced (no standarized technique). After cardiac surgery, at the beginning of skin suture, inhaled anesthetics were discontinued and after successful spontaneous breathing test, the patient was extubated in the operating room. PaO2 y PaCO2 was evaluated at the arrival to the ICU. Visual analogue scale (VAS) was applied every 6 hours to measure pain level. At the end of the first day opiod dose, PONV and ventilation support (invasive and noinvasive) was registered...(AU)

Nerve growth factor induces the expression of chaperone protein calreticulin in human epithelial ovarian cells.

Epithelial ovarian cancer is highly angiogenic and high expression of Nerve Growth Factor (NGF), a proangiogenic protein. Calreticulin is a multifunctional protein with anti-angiogenic properties and its translocation to the tumor cell membrane promotes recognition and engulfment by dendritic cells. The aim of this work was to evaluate calreticulin expression in human normal ovaries, benign and borderline tumors, and epithelial ovarian cancer samples and to evaluate whether NGF regulates calreticulin expression in human ovarian surface epithelium and in epithelial ovarian cancer cell lines. Calreticulin mRNA and protein levels were analyzed using RT-PCR, Western blot and immunohistochemistry in 67 human ovarian samples obtained from our Institution. Calreticulin expression induced by NGF stimulation in cell lines was evaluated using RT-PCR, Western blot and immunocytochemistry. We found a significant increase of calreticulin mRNA levels in epithelial ovarian cancer samples as compared to normal ovaries, benign tumors, and borderline tumors. Calreticulin protein levels, evaluated by Western blot, were also increased in epithelial ovarian cancer with respect to benign and borderline tumors. When HOSE and A2780 cell lines were stimulated with Nerve Growth Factor, we found an increase in calreticulin protein levels compared to controls. This effect was reverted by GW441756, a TRKA specific inhibitor. These results suggest that NGF regulates calreticulin protein levels in epithelial ovarian cells through TRKA receptor activation.

Autotrasplante cardiaco en el tratamiento quirúrgico de tumores cardiacos primarios: Caso clínico / Heart autotrasplantation for the treatment of a rhabdomyosarcoma of the left ventricle: Report of one case

Background: Heart autotrasplantation is an exceptional surgical technique used in the treatment of uncontrolled cardiac arrhythmias and primary unresectable cardiac tumors. We report a 28-year-old male with a rhabdomyosarcoma of the left ventricle, localized in the lateral and posterior wall, which involved the mitral valve and circumflex artery. After a complete study ruling out dissemination of the tumor, the patient was operated. Surgical exploration determined the unresectabilility of the tumor with the heart in situ. Therefore, the heart was explanted, preserving the right atrium and coronary sinus for re-implantation. Fifty percent of the mitral valve and the circumflex artery from its origin, were resected due to tumor infiltration. The heart was reconstructed with bovine pericardium and a mechanical valve was implanted in the mitral position. Afterward, the heart was implanted again following the same sequence as in bicaval transplantation, followed by a double bypass grafting to the distal circumflex territory. The patient had no significant complications and after nine months of follow up, there was no evidence of local recurrence. In the fourth postoperative month, a subcutaneous mass in the left thigh that was considered a metastasis without histological confirmation appeared. The lesion disappeared with radio and chemotherapy.

Nerve growth factor stimulates cellular proliferation of human epithelial ovarian cancer.

Due to its ability to induce vascular endothelial growth factor expression and proliferation, migration, and vasculogenesis of endothelial cells, nerve growth factor (NGF) has been considered as an angiogenic factor in epithelial ovarian cancer (EOC). In this work, we evaluated the angiogenic and proliferative mRNA expression profiles of EOC and addressed the responsiveness of EOC explants to NGF stimulation. Twenty EOC samples were obtained from Obstetrics and Gynecology Department, University of Chile's Clinical Hospital. Global gene expression profiles of selected poorly differentiated serous EOC samples were obtained with DNA oligonucleotide microarrays. In addition, EOC explants were subjected to NGF stimulation and levels of p-AKT, BAX, BCL2, Ki-67, c-MYC, and FOXL2 proteins were determined by immunohistochemistry. Results showed that mRNAs coding for specific transcriptional regulators and antiapoptotic components of the NGF signaling pathway were upregulated in EOC cells. At the protein level, key members of the NGF pathway including p-AKT, BCL2/BAX, Ki-67, and c-MYC were found increased, while FOXL2 was decreased in response to NGF stimulation. These findings strongly suggest that NGF stimulates cellular proliferation of human EOC.

[Heart autotransplantation for the treatment of a rhabdomyosarcoma of the left ventricle. Report of one case]. / Autotrasplante cardiaco en el tratamiento quirúrgico de tumores cardiacos primarios. Caso clínico.

Heart autotransplantation is an exceptional surgical technique used in the treatment of uncontrolled cardiac arrhythmias and primary unresectable cardiac tumors. We report a 28-year-old male with a rhabdomyosarcoma of the left ventricle, localized in the lateral and posterior wall, which involved the mitral valve and circumflex artery. After a complete study ruling out dissemination of the tumor, the patient was operated. Surgical exploration determined the unresectability of the tumor with the heart in situ. Therefore, the heart was explanted, preserving the right atrium and coronary sinus for re-implantation. Fifty percent of the mitral valve and the circumflex artery from its origin, were resected due to tumor infiltration. The heart was reconstructed with bovine pericardium and a mechanical valve was implanted in the mitral position. Afterward, the heart was implanted again following the same sequence as in bicaval transplantation, followed by a double bypass grafting to the distal circumflex territory. The patient had no significant complications and after nine months of follow up, there was no evidence of local recurrence. In the fourth postoperative month, a subcutaneous mass in the left thigh that was considered a metastasis without histological confirmation appeared. The lesion disappeared with radio and chemotherapy.

Disección aórtica tipo A: Resultados operatorios y seguimiento a mediano plazo / Surgical results among 100 patients with type A aortic dissection: Retrospective review

Background: Tipe A aortic dissection involves the ascending aorta and has high mortality rates without surgical treatment. Aim: To report the results of surgical treatment of type A aortic dissection. Material and Methods: Retrospective review of medical records of 100 patients aged 17 to 78 years (73 percent males) operated between January 2000 and August 2008, for type A aortic dissection. Follow up was performed with telephone interviews and review of national death records. Results: Eighty three percent of patients had an acute dissection. Operative mortality was 27 and 20 percent for patients with acute and chronic dissection, respectively. Mortality was 50 percent among patients aged 70 years or more, compared with 21 percent among their younger counterparts, The most common complication was bleeding that required a new surgical procedure in 18 percent of patients. Actuarial survival was 70 percent at five years. Cardiovascular problems caused the death of two of the nine patients that died during follow up. Conclusions: Surgical mortality among patients with type A aortic dissection was higher among patients with acute episodes and those aged 70 years or more.

Disección aórtica y síndrome de Marfán en el embarazo: A propósito de un caso clínico / Acute type A aortic dissection in pregnant patient with Marfan syndrome: Report of one case

Aortic dissection is a life-threatening disease that requires immediate surgical intervention. Marfan syndrome is a hereditary disease with an autosomaldominant transmission, which affects the connective tissue, with skeletal, cardiovascular and ocular involvement. It is one of the most prevalent connective tissue disorders, presenting a risk of aortic dissection of approximately 1% even without dilatation of the aorta. When dissectionoccurs during pregnancy and requires surgical intervention (type A dissection), maternal mortality is high (20%-30%). We report a 38 year-old woman with Marfan syndrome that hadan acute type A aortic dissection and severe aortic regurgitation at 37 weeks of gestation. The patient underwent a cesarean section and delivered a healthy baby. Afterwards, aortic valverepair and ascending aortic replacement was successfully performed under circulatory arrest with deep hypothermia. Additionally mitral valve repair for degenerative disease with posterior, autologous pericardium mitral valve ring was performed.

Incisiones transversas en oncología ginecológica / Transverse incisions in gynecologic oncology

OBJETIVO: Determinar si el tipo de laparotomía influye en la etapificación de pacientes con cáncer de cuerpo y cuello uterino. MÉTODO: Se revisaron todas las fichas clínicas de pacientes con cáncer de cuerpo y cuello uterino que fueron operadas en el Hospital Clínico de la Universidad de Chile y el Hospital Clínico de la Fuerza Aérea Chilena, entre enero de 1999 y mayo de 2005. Se recopiló la siguiente información: tipo de laparotomía, índice de masa corporal (IMC), comorbilidades médicas, tiempo operatorio, histología, número total y distribución de linfonodos, pérdida sanguínea, complicaciones, duración de la cirugía y hospitalización. Se aplicaron análisis estadísticos con t student y c². RESULTADOS: Se identificaron 51 pacientes. Se usó laparotomía media (LM) y transversa (LT) en 16 (31%) y 35 (69%) de las pacientes, respectivamente. No hubo diferencias significativas en índice de masa corporal, estadio FIGO, histología, comorbilidades, estimación de pérdida sanguínea ni complicaciones intra o post operatorias entre el grupo de LM y LT. Se encontraron diferencias significativas en pacientes con IMC 25 sometidas a laparotomías verticales comparadas con las transversas, donde ocurrió mayor sangrado intraoperatorio, se recolectó mayor número de ganglios para-aórticos y tuvieron hospitalizaciones más prolongadas. CONCLUSIONES: La etapificación quirúrgica de pacientes con cáncer de cérvix o cuerpo uterino se puede realizar adecuadamente a través de incisiones transversas, sin mayor morbilidad. Previa adecuada selección, pacientes con cánceres cervical y uterino pueden beneficiarse de las ventajas ya descritas para las laparotomías transversas.

OBJECTIVE: To determine if the type of abdominal incision influences the adequacy of surgical staging in patients with uterine and cervical cancer. METHODS: A retrospective review of all uterine and cervical cancer patients operated on by the same surgeon at the Universidad de Chile Clinical Hospital and the Chilean Air Force Clinical Hospital between January 1, 1999, and May 1, 2005, is presented. Data on type of incision, body mass index (BMI), medical comorbilities, histology, total number and distribution of lymph nodes, estimated blood loss, complications, length of surgery and hospital stay were abstracted. Statistical analysis with two-tailed Student t test and c² were performed. RESULTS: 51 patients were identified. A vertical incision (VI) was used in 16 (31%) while 35 (69%) received a transverse incision (TI). There were no statistically significant differences in BMI, FIGO stage, histology, comorbilities, estimated blood loss or intraoperative and postoperative complications between the VI and TI groups. Compared patients with BMI 25, VI was associated with significantly more intraoperative blood loss, number of para-aortic lymph nodes harvested and length of hospital stay. CONCLUSION: Comprehensive surgical staging for uterine and cervical cancer can be adequately performed through a TI without greater morbidity. After appropriate selection, patients with uterine and cervical cancer can benefit from the inherent benefits previously described for TI.

Precisión diagnóstica y riesgo de implante secundario en el estudio de carcinoma endometrial por histeroscopía / Diagnostic accuracy and risk of secondary implant in the study of endometrial carcinoma by hysteroscopy

Adenocarcinoma of the endometrium is the most common malignancy of the female genital tract in developed countries. Hysteroscopy has turned into a widely used procedure to improve curettage-biopsy sensibility. Uncertainties have arisen about the hysteroscopic endometrial carcinoma diagnostic sensibility its safety due to the possibility of peritoneal seeding. A MEDLINE and Cochrane Library search was performed. We found that the hysteroscopic diagnosis has low incidence of complications and high diagnostic rate for endometrial carcinoma. There are cytological, case reports and retrospectives studies showing both an increase and also non-increase of seeding risk of cancer cells.

Immunotherapeutic potential of DISC-HSV and OX40L in cancer.

Several vectors, viral and bacterial, have been developed over the past few years for means of generating an effective antitumor immune response. We have developed and studied a "model for immunotherapy" using a viral vector disabled infectious single cycle-herpes simplex virus (DISC-HSV), which efficiently transduces various tumor cell lines and offers a useful vehicle for the further development of cell-based vaccines. The immunotherapeutic potential of DISC-HSV encoding granulocyte macrophage colony stimulating factor (GM-CSF) was demonstrated in a number of murine carcinoma models, leading to complete regression of well-established tumors in up to 70% of the mice. Moreover, the therapeutic potential of DISC-HSV-GM-CSF was significantly enhanced when used in combination therapy with either OX40L or dendritic cells (DC), even in a poorly immunogenic tumor model. The ability of this vector to accept large gene inserts, its good safety profile, its ability to undergo only a single round of infection, the inherent viral immunostimulatory properties and its ability to infect various tumor cell lines efficiently, make DISC-HSV an ideal candidate vector for immunotherapy. The DISC- CT-26 tumor model was used to investigate the mechanisms associated with immunotherapy induced tumor rejection. Although CTL induction, was positively correlated with regression, MHC class I down regulation and accumulation of immature Gr1+ myeloid cells were shown to be the main immuno-suppressor mechanisms operating against regression and associated with progressive tumor growth. The CTL response was associated with the immuno-dominant AH-1 peptide of the retroviral glycoprotein gp70. This model of immunotherapy has provided an opportunity to dissect further the immunological events associated with tumor-rejection and escape. Since other antigens may be important in initiating tumor rejection, we have investigated the expression of MTA-1, an antigen that appears to be expressed widely in human and murine tumors. The immunogenicity of MTA-1 was studied and its potential as a tumor rejection antigen is under investigation.

Immunity to tumour antigens.

During the last decade, a large number of human tumour antigens have been identified. These antigens are classified as tumour-specific shared antigens, tissue-specific differentiation antigens, overexpressed antigens, tumour antigens resulting from mutations, viral antigens and fusion proteins. Antigens recognised by effectors of immune system are potential targets for antigen-specific cancer immunotherapy. However, most tumour antigens are self-proteins and are generally of low immunogenicity and the immune response elicited towards these tumour antigens is not always effective. Strategies to induce and enhance the tumour antigen-specific response are needed. This review will summarise the approaches to discovery of tumour antigens, the current status of tumour antigens, and their potential application to cancer treatment.

Regulation of CTL responses to MHC-restricted class I peptide of the gp70 tumour antigen by splenic parenchymal CD4+ T cells in mice failing immunotherapy with DISC-mGM-CSF.

Direct intratumour injection of the disabled infectious single-cycle-herpes simplex virus-encoding murine granulocyte/macrophage colony-stimulating factor (DISC-HSV-mGM-CSF) into established colon carcinoma CT26 tumours induced complete tumour rejection in up to 70% of treated animals (regressors), while the remaining mice developed progressive tumours (progressors). This murine Balb/c model was used to dissect the cellular mechanisms involved in tumour regression or progression following immunotherapy. CTLs were generated by coculturing lymphocytes and parenchymal cells from the same spleens of individual regressor or progressor animals in the presence of the relevant AH-1 peptide derived from the gp70 tumour-associated antigens expressed by CT26 tumours. Tumour regression was correlated with potent CTL responses, spleen weight and cytokine (IFN-gamma) production. Conversely, progressor splenocytes exhibited weak to no CTL activity and poor IFN-gamma production, concomitant with the presence of a suppressor cell population in the progressor splenic parenchymal cell fraction. Further fractionation of this parenchymal subpopulation demonstrated that cells inhibitory to the activation of AH-1-specific CTLs, restimulated in vitro with peptide, were present in the nonadherent parenchymal fraction. In vitro depletion of progressor parenchymal CD3+/CD4+ T cells restored the CTL response of the cocultured splenocytes (regressor lymphocytes and progressor parenchymal cells) and decreased the production of IL-10, suggesting that CD3+CD4+ T lymphocytes present in the parenchymal fraction regulated the CTL response to AH-1. We examined the cellular responses associated with tumour rejection and progression, identifying regulatory pathways associated with failure to respond to immunotherapy.

Peptide immunisation of HLA-DR-transgenic mice permits the identification of a novel HLA-DRbeta1*0101- and HLA-DRbeta1*0401-restricted epitope from p53.

Because of the central role of CD4(+) T cells in antitumour immunity, the identification of the MHC class II-restricted peptides to which CD4(+) T cells respond has become a priority of tumour immunologists. Here, we describe a strategy permitting us to rapidly determine the immunogenicity of candidate HLA-DR-restricted peptides using peptide immunisation of HLA-DR-transgenic mice, followed by assessment of the response in vitro. This strategy was successfully applied to the reported haemaglutinin influenza peptide HA(307-319), and then extended to three candidate HLA-DR-restricted p53 peptides predicted by the evidence-based algorithm SYFPEITHI to bind to HLA-DRbeta1*0101 (HLA-DR1) and HLA-DRbeta1*0401 (HLA-DR4) molecules. One of these peptides, p53(108-122), consistently induced responses in HLA-DR1- and in HLA-DR4-transgenic mice. Moreover, this peptide was naturally processed by dendritic cells (DCs), and induced specific proliferation in the splenocytes of mice immunised with p53 cDNA, demonstrating that immune responses could be naturally mounted to the peptide. Furthermore, p53(108-122) peptide was also immunogenic in HLA-DR1 and HLA-DR4 healthy donors. Thus, the use of this transgenic model permitted the identification of a novel HLA-DR-restricted epitope from p53 and constitutes an attractive approach for the rapid identification of novel immunogenic MHC class II-restricted peptides from tumour antigens, which can ultimately be incorporated in immunotherapeutic protocols.