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Rationale: The tumor microenvironment (TME) and its multifaceted interactions with cancer cells are major targets for cancer treatment. Single-cell technologies have brought major insights into the TME, but the resulting complexity often precludes conclusions on function. Methods: We combined single-cell RNA sequencing and spatial transcriptomic data to explore the relationship between different cancer-associated fibroblast (CAF) populations and immune cell exclusion in breast tumors. The significance of the findings was then evaluated in a cohort of tumors (N=75) from breast cancer patients using immunohistochemistry analysis. Results: Our data show for the first time the degree of spatial organization of different CAF populations in breast cancer. We found that IL-iCAFs, Detox-iCAFs, and IFNγ-iCAFs tended to cluster together, while Wound-myCAFs, TGFβ-myCAFs, and ECM-myCAFs formed another group that overlapped with elevated TGF-β signaling. Differential gene expression analysis of areas with CD8 + T-cell infiltration/exclusion within the TGF-β signaling-rich zones identified elastin microfibrillar interface protein 1 ( EMILIN1 ) as a top modulated gene. EMILIN1 , a TGF-β inhibitor, was upregulated in IFNγ-iCAFs directly modulating TGFβ immunosuppressive function. Histological analysis of 75 breast cancer samples confirmed that high EMILIN1 expression in the tumor margins was related to high CD8 + T-cell infiltration, consistent with our spatial gene expression analysis. High EMILIN1 expression was also associated with better prognosis of patients with breast cancer, underscoring its functional significance for the recruitment of cytotoxic T cells into the tumor area. Conclusion: Our data show that correlating TGF-β signaling to a CAF subpopulation is not enough because proteins with TGF-β-modulating activity originating from other CAF subpopulations can alter its activity. Therefore, therapeutic targeting should remain focused on biological processes rather than on specific CAF subtypes., Competing Interests: Competing Interests: The authors have declared that no competing interest exists., (© The author(s).)

The study of cellular networks mediated by ligand-receptor interactions has attracted much attention recently owing to single-cell omics. However, rich collections of bulk data accompanied with clinical information exists and continue to be generated with no equivalent in single-cell so far. In parallel, spatial transcriptomic (ST) analyses represent a revolutionary tool in biology. A large number of ST projects rely on multicellular resolution, for instance the Visium™ platform, where several cells are analyzed at each location, thus producing localized bulk data. Here, we describe BulkSignalR, a R package to infer ligand-receptor networks from bulk data. BulkSignalR integrates ligand-receptor interactions with downstream pathways to estimate statistical significance. A range of visualization methods complement the statistics, including functions dedicated to spatial data. We demonstrate BulkSignalR relevance using different datasets, including new Visium liver metastasis ST data, with experimental validation of protein colocalization. A comparison with other ST packages shows the significantly higher quality of BulkSignalR inferences. BulkSignalR can be applied to any species thanks to its built-in generic ortholog mapping functionality., (© The Author(s) 2023. Published by Oxford University Press on behalf of Nucleic Acids Research.)

Rationale: Patients with colorectal cancer die mainly due to liver metastases (CRC-LM). Although the tumor microenvironment (TME) plays an important role in tumor development and therapeutic response, our understanding of the individual TME components, especially cancer-associated fibroblasts (CAFs), remains limited. Methods: We analyzed CRC-LM CAFs and cancer cells by single-cell transcriptomics and used bioinformatics for data analysis and integration with related available single-cell and bulk transcriptomic datasets. We validated key findings by RT-qPCR, western blotting, and immunofluorescence. Results: By single-cell transcriptomic analysis of 4,397 CAFs from six CRC-LM samples, we identified two main CAF populations, contractile CAFs and extracellular matrix (ECM)-remodeling/pro-angiogenic CAFs, and four subpopulations with distinct phenotypes. We found that ECM-remodeling/pro-angiogenic CAFs derive from portal resident fibroblasts. They associate with areas of strong desmoplastic reaction and Wnt signaling in low-proliferating tumor cells engulfed in a stiff extracellular matrix. By integrating public single-cell primary liver tumor data, we propose a model to explain how different liver malignancies recruit CAFs of different origins to this organ. Lastly, we found that LTBP2 plays an important role in modulating collagen biosynthesis, ECM organization, and adhesion pathways. We developed fully human antibodies against LTBP2 that depleted LTBP2+ CAFs in vitro . Conclusion: This study complements recent reports on CRC-LM CAF heterogeneity at the single-cell resolution. The number of sequenced CAFs was more than one order of magnitude larger compared to existing data. LTBP2 targeting by antibodies might create opportunities to deplete ECM-remodeling CAFs in CRC-LMs. This might be combined with other therapies, e.g., anti-angiogenic compounds as already done in CRC. Moreover, we showed that in intrahepatic cholangiocarcinoma, in which ECM-remodeling CAF proportion is similar to that of CRC-LM, several genes expressed by ECM-remodeling CAFs, such as LTBP2 , were associated with survival., Competing Interests: Competing Interests: The authors have declared that no competing interest exists., (© The author(s).)

Background: Biomarkers are urgently needed for pancreatic ductal adenocarcinoma (PDAC). Endoscopic ultrasound-guided fine-needle aspiration (EUS-FNA) is the cornerstone for diagnosing PDAC. We developed a method for discovery of PDAC biomarkers using the discarded EUS-FNA liquid., Methods: This retrospective study included 58 patients with suspected pancreatic lesions who underwent EUS-FNA. Protein extracts from EUS-FNA liquid were analyzed by mass spectrometry. Proteomic and clinical data were modeled by supervised statistical learning to identify protein markers and clinical variables that distinguish PDAC., Results: Statistical modeling revealed a protein signature for PDAC screening that achieved high sensitivity and specificity (0.92, 95 % confidence interval [CI] 0.79-0.98, and 0.85, 95 %CI 0.67-0.93, respectively). We also developed a protein signature score (PSS) to guide PDAC diagnosis. In combination with patient age, the PSS achieved 100 % certainty in correctly identifying PDAC patients > 54 years. In addition, 3 /4 inconclusive EUS-FNA biopsies were correctly identified using PSS., Conclusions: EUS-FNA-derived fluid is a rich source of PDAC proteins with biomarker potential. The PSS requires further validation and verification of the feasibility of measuring these proteins in patient sera., Competing Interests: The authors declare that they have no conflict of interest., (Thieme. All rights reserved.)

Cancer-associated fibroblasts (CAF) are important constituents of the tumor microenvironment (TME) and are major drivers of tumorigenesis. Yet, therapies aiming at eliminating CAF have failed to cure patients. This setback has raised questions regarding whether CAF exclusively favour cancer progression, or if they may also assume tumor-suppressor functions. In the present study, we used proteomics and single cell RNA-sequencing analysis to examine the CAF landscape in hepatocellular carcinoma (HCC). We thereby unveil three major CAF populations in HCC, one of which specifically expressing the prolargin protein. This CAF subpopulation (further termed as CAF_Port) shared a strong transcriptomic signature with portal liver fibroblasts. We further show that CAF_Port deposit prolargin in the TME and that its levels are lower in tumors as compared to the peritumoral region. Mechanistically, aggressive cancer cells degraded prolargin using matrix metalloprotease activity. Survival analysis of 188 patients revealed that high prolargin protein levels correlate with good patient outcome (HR = 0.37; p = 0.01). In vivo, co-injection of cancer cells with fibroblasts silenced for prolargin, led to faster tumor development (5-fold; p = 0.01), mainly due to stronger angiogenesis. Using protein-protein interaction study and structural modelling, we further demonstrate that prolargin binds and inhibits the activity of several pro-agiogenic proteins, including hepatocyte and fibroblast growth factors. In conclusion, prolargin is angiogenesis modulator and CAF-derived tumor suppressor in HCC. Stabilizing prolargin levels in the CAF_Port subpopulation may revert their tumor-antagonizing properties, warranting exploration in further pre-clinical studies., (© 2022. The Author(s), under exclusive licence to Springer Nature Limited.)

γδ T cells contribute to the immune response against many cancers, notably through their powerful effector functions that lead to the elimination of tumor cells and the recruitment of other immune cells. However, their presence in the tumor microenvironment has been associated with poor prognosis in breast, colon, and pancreatic cancer, suggesting that γδ T cells may also display pro-tumor activities. Here, we identified in blood from healthy donors a subpopulation of Vδ1T cells that represents around 20% of the whole Vδ1 population, expresses CD73, and displays immunosuppressive phenotype and functions (i.e., production of immunosuppressive molecules, such as IL-10, adenosine, and the chemotactic factor IL-8, and inhibition of αβ T cell proliferation). We then found that in human breast tumors, γδ T cells were present particularly in late stage breast cancer samples, and that ∼20% of tumor-infiltrating γδ T cells expressed CD73. Taken together, these results suggest that regulatory γδ T cells are present in the breast cancer microenvironment and may display immunosuppressive functions through the production of immunosuppressive molecules, such as IL-10, IL-8, and adenosine, thus promoting tumor growth., (©2020 Society for Leukocyte Biology.)

Background: Circulating mesenchymal stem cells contribute to bone repair. Their incorporation in fracture callus is correlated to their bioavailability. In addition, Granulocyte-colony stimulating factor induces the release of vascular and mesenchymal progenitors. We hypothesized that this glycoprotein stimulates fracture healing, and analyzed the effects of its administration at low doses on bone healing., Methods: 27 adult male Sprague-Dawley rats underwent mid-femur osteotomy stabilized by centromedullar pinning. In a post (pre) operative group, rats were subcutaneously injected with 5 μg/kg per day of Granulocyte-colony stimulating factor for 5 days after (before) surgery. In a control group, rats were injected with saline solution for 5 days immediately after surgery. A radiographic consolidation score was calculated. At day 35, femurs were studied histologically and underwent biomechanical tests., Findings: 5 weeks after surgery, mean radiographic scores were significantly higher in the Preop group 7.75 (SD 0.42) and in the Postop group 7.67 (SD 0.52) than in the control group 6.75 (SD 0.69). Biomechanical tests showed femur stiffness to be more than three times higher in both the Preop 109.24 N/mm (SD 51.86) and Postop groups 100.05 N/mm (SD 60.24) than in control 32.01 N/mm (SD 15.78). Mean maximal failure force was twice as high in the Preop group 68.66 N (SD 27.78) as in the control group 34.21 N (SD 11.79). Histological results indicated a later consolidation process in control than in treated groups., Interpretation: Granulocyte-colony stimulating factor injections strongly stimulated early femur fracture healing, indicating its potential utility in human clinical situations such as programmed osteotomy and fracture., (Copyright © 2018. Published by Elsevier Ltd.)

Advanced Epithelial Ovarian Cancer (EOC) patients frequently relapse by 24 months and develop resistant disease. Research on EOC therapies relies on cancer cell lines established decades ago making Patient Derived Xenografts (PDX) attractive models, because they are faithful representations of the original tumor. We established 35 ovarian cancer PDXs resulting from the original graft of 77 EOC samples onto immuno-compromised mice. PDXs covered the diversity of EOC histotypes and graft take was correlated with early patient death. Fourteen PDXs were characterized at the genetic and histological levels. PDXs reproduced phenotypic features of the ovarian tumors of origin and conserved the principal characteristics of the original copy number change (CNC) profiles over several passages. However, CNC fluctuations in specific subregions comparing the original tumor and the PDXs indicated the oligoclonal nature of the original tumors. Detailed analysis by CGH, FISH and exome sequencing of one case, for which several tumor nodules were sampled and grafted, revealed that PDXs globally maintained an oligoclonal structure. No overgrowth of a particular subclone present in the original tumor was observed in the PDXs. This suggested that xenotransplantation of ovarian tumors and growth as PDX preserved at least in part the clonal diversity of the original tumor. We believe our data reinforce the potential of PDX as exquisite tools in pre-clinical assays.

Introduction: The prevention or the management of digestive fistulae may be performed by using an external wrap of collagen of animal origin. To evaluate this treatment, an experimental study creating a hole in the colon of pig covered by a resorbable collagen belt was performed. Results are very interesting and collagen wrap is very well tolerated by the colon wall., Background: Digestive perforations, whether colorectal, jejunal, esophageal, or biliodigestive, are common emergency situations and can threaten the patient's condition or extend their hospital stay. The evolution of biomaterials of animal origin, and the biocompatibility proven after some human surgical procedures, have led our team to propose an experimental study in a pig model to treat colic perforation by positioning a resorbable bilayer collagen band of bovine origin over the area of an experimental hole., Materials/methods: A first group of 10 pigs was operated upon, and a 1 cm2 hole was experimentally created in the distal part of the colon. Then, a belt of resorbable collagen sponge joined to a collagen film, from bovine origin, was placed and fixed around the outer part of the colon to cover the fistula without closing the hole by sutures. After an average of two weeks, all the animals were sacrificed. The abdominal cavity was examined in a macroscopic and microscopic manner. A second group of 10 pigs was tested under a different protocol to assess the efficiency of the bowel wrap prosthesis in a septic field., Results: In the first group of pigs, there were no complications during the procedures. The mortality rate was zero during this period. No pig was operated on urgently to manage an acute complication. The complication rate was 10% due to one wound infection. The macroscopic examinations of the explanted colon articles didn't find any stricture under the prosthesis location for the 10 pigs. Local smooth adhesions were noted in 7 pigs (70%). Among the second group of pigs, the mortality rate was 10% due to a myocardial infarction during the period of peritonitis. No pig was operated on urgently to manage an acute complication. The complication rate was 20% due to 2 wound infections. The macroscopic examination of the explanted colon articles found one case of stricture under the prosthesis location (10%). Local smooth adhesions were noted in 7 pigs (70%). No histologic rejection was noted during the anatomopathologic tests for all pigs., Conclusion: The use of bovine collagen bilayer prosthesis in digestive surgery may prove to be safe and effective to treat digestive leakage. It may be feasible to use this type of biomaterial to prevent fistula of the digestive tract, including anastomotic. A prospective trial would need to be performed to complete this research to give the surgeons an opportunity to improve treatment in many digestive procedures.

Introduction: The purpose of the study was to compare the efficacy on rat skull defects of two bone growth factors derived from the GDF-5 family., Material and Methods: The study was conducted on 17 adult Wistar rats. On each animal, two symmetrical 6-mm wide, full-thickness, skull defects were carried out in the parietal regions. In 15 out of 17 animals, both experimental defects were filled by the implants. In the group I (n=2), both defects were left empty for control. The 15 other rats were divided into 3 groups: In group II (n=5), a collagen sponge was implanted. In group III (n=5), a collagen sponge impregnated with rhGDF-5 (the genuine dimeric form) was implanted. In group IV (n=5), a collagen sponge impregnated with rhGDF-5C465A (a monomeric form of GDF-5) was implanted. All animals were sacrificed at 8 weeks. The harvested specimens were processed for contact radiography and standard histological examination. The quantitative results were assessed with a semi-quantitative histological scoring system., Results: One animal in the group II was excluded because it died of unknown reasons. In group I, no bone healing was observed in the defects. In group II, no bone healing was observed in 4 out of 10 defects, and partial bone healing was observed in 5 out of 10 defects. In group III, partial bone healing was also observed in 3 out of 8 defects and complete bone healing in 4 out of 8 defects. In group IV, partial bone healing was observed in 8 out of 10 defects and complete bone healing in 2 out of 10 defects., Conclusion: Bone healing was improved in all treated groups. Further studies are necessary to determine the optimal formulation of these composite implants.

Our retrospective study analyzes various factors to evaluate the risk of invasion of the not sentinel node when the sentinel node biopsy is positive in the infiltrated breast cancers. We compared in single varied then multivaried analysis, various parameters between two groups: positive not sentinel nodes and negative not sentinel nodes among 180 cases of positive sentinel node biopsy between 2001 and 2004. At the time of the single varied analysis, seem to be risk factors of non sentinel node involvement: the histopronostic SBRIII rank, positive a HER2neu status, the presence of extracapsulal node extension and infiltration of the sentinel node by a macrometastasis. The tumoral embol, the absence of hormonal receivers, a tumoral size > 10 mm and the number of sentinel node taken appear at the limit of the significativity. In multivaried analysis, SBRIII rank and the presence of an extracapsular node extension remain related to non sentinel node involvement. The histological type, association with a CIS, the size of the sentinel nodes, the number of positive sentinel nodes and the year of surgery are nonsignificant. Additional axillairy clearing out at the time of a positive node sentinel biopsy should be discussed according to different criteria determined by a precise histological analysis.

Background: To maintain the long-term effects of a gastric bariatric operation, bands are often placed to control the restriction. Erosion into the gastric wall by these devices remains a problem. A soft resiliant prosthesis of animal origin, constituted by a network of non-absorbable collagen fibres, may be a solution to this problem. This study assessed, in a porcine model, the histological reaction of the gastric wall following apposition of a band of porcine collagen (Pelvicol, Bard)., Methods: 15 female pigs weighing on average 21 kg underwent vertical banded gastroplasty (VBG). Stoma control was achieved with a band of porcine collagen (2 cm wide, 7 cm long and 2 mm thick). The pigs were sacrificed 1 month after VBG, and histological analysis was performed at a macroscopic and microscopic level., Results: There was no peri-operative death, although 2 pigs died in the postoperative period (the first case developed a bowel fistula and sepsis, and the second pig died of unrelated causes). There were 2 additional morbidities (gastric fistula on the linear staple-line away from the Pelvicol band) that led to an early euthanasia of 2 pigs. Post-mortem macroscopic analyses in the remaining 11 pigs did not reveal migration of the device, and there was no tissue reaction on postoperative microscopic analyses. 10 of the pigs had lost weight at 1 month, averaging 3.42 kg., Conclusion: Porcine collagen appears to be an effective and safe alternative to the current methods of control of pouch outlet. The flexibility and homogeneity of this prosthesis may be useful to limit the risk of erosion of the gastric wall. Although these properties have been assessed in pelvic operations in humans, this work needs to be studied in a prospective long-term study in humans.

Anti-Müllerian hormone (AMH) [also called Müllerian inhibiting substance (MIS)] is a member of the transforming growth factor-beta family. AMH and its type II receptor (AMHR-II) are involved in the regression of the Müllerian ducts in the male embryo, and in gonadal functions in the adult. AMH is also known to be a marker of granulosa and Sertoli cell tumours. We selected a high-affinity monoclonal antibody, mAb 12G4, specific for human AMHR-II (hAMHR-II), by FACS analysis, Western blotting and immunohistochemical staining of a hAMHR-II-transfected CHO (Chinese hamster ovary) cell line, normal adult testicular tissue and granulosa cell tumours. Using peptide array screening, we identified the binding sequences of mAb 12G4 and AMH on the receptor. Identification of Asp53 and Ala55 as critical residues in the DRAQVEM minimal epitopic sequence of mAb 12G4 definitively accounted for the lack of cross-reactivity with the murine receptor, in which there is a glycine residue in place of an aspartic acid residue. In a structural model, the AMH-binding interface was mapped to the concave side of hAMHR-II, whereas the mAb 12G4-binding site was located on the convex side. mAb 12G4, the first mAb to be raised against hAMHR-II, therefore has unique properties that could make it a valuable tool for the immunotargeting of tumours expressing this receptor.

Carcinoembryonic antigen (CEA) and ErbB-2 are expressed in about 50 and 30% of breast cancers, respectively. We hypothesised that targeting of these two antigens by a bispecific antibody (BAb) might provide efficient tumour uptake and prolonged tumour residence time. In the present study, we first studied the expression of CEA and ErbB-2 on primary breast tumours screened by immunohistochemistry. Of 106 primary breast cancers, 69 (65%) were positive for CEA, 20 (19%) were positive for ErbB-2, and 13 (12%) expressed both antigens. We then prepared and evaluated a BAb directed against CEA and ErbB-2. Using BIACORE technology, we showed that the BAb recognised both CEA and ErbB-2 with affinities of 0.9 x 10 and 0.8 x 10 M(-1), respectively. In vivo, BAb tumour localisation was compared with that of its parental homodimeric F(ab')(2)-ORTHO-phenylene- dimaleimide (PDM) fragments. Uptake of (125)I-BAb was lower than that of (131)I-35A7F(ab')(2)-PDM in LS174T tumours, used as a model of CEA expressing tumours, and was similar to that of (131)I-FWP51 F(ab')(2)-PDM in SKOv3 tumours, used as a model of ErbB-2 expressing tumours. In a double-positive model, the SKOv3-CEA-1B9 tumour, BAb showed a similar uptake to that of 35A7 F(ab')(2)-PDM and we demonstrated that, although BAb had double specificity, it internalised as a homodimeric anti-ErbB-2 antibody. BAb showed a greater uptake than that of FWP51 F(ab')(2)-PDM and this difference was even more important 72 h after injection with an uptake of 7.3 +/- 2.1 vs. 1.4 +/- 0.5% of the injected dose per gram of tissue. The results obtained with the BAb in the double-positive tumour-bearing nude mice suggest that targeting two distinct tumour-associated antigens on the same cell could improve tumour localisation., (Copyright 2002 S. Karger AG, Basel)

The aim of our study was to assess the technique of immunophotodetection (IPD) in intraoperative situations in an experimental model and to determine its capacity to detect very small tumor masses. IPD is a recent technology involving fluorescent dye-labeled monoclonal antibodies (MAbs) directed against tumor-associated antigens. Up to now, no intraoperative device for IPD has been developed, and limits of detection of the technique are unknown. MAb-dye conjugates were prepared using the anti-carcinoembryonic antigen MAb 35A7 labeled with indocyanine and (125)I. Time-dependent (6, 12, 24, 48, and 96 h post i.v. injection) and dose-dependent (10, 40, and 100 microg of conjugate) biodistribution studies were performed in nude mice bearing an LS174T peritoneal carcinomatosis demonstrating high tumor uptake (up to 21% of the injected dose/g of tumor 48 h postinjection). Intraoperative IPD was studied, using a newly developed device, in 16 mice 48 h after i.v. injection of 40 microg of the (125)I-MAb 35A7-indocyanine conjugate. The fluorescent status of 333 biopsies was compared with their histological analysis. Sensitivity was 90.7%, specificity was 97.2%, the positive predictive value was 94.7%, and the negative predictive value was 94.9%. Detection of very small nodules (<1 mg in weight or <1 mm in diameter) was possible. However, we observed a decrease in sensitivity as a function of tumor mass: 100% for nodules >10 mg versus 78% for nodules < or =1 mg. These experiments demonstrate that intraoperative IPD is easy to use and associated with high sensitivity and specificity, even for low tumor masses. On the basis of these encouraging results, intraoperative IPD should be assessed in a clinical study.

Eggshell has been recently introduced as a bone substitute candidate in reconstructive surgery. The aim of this experimental study study was to determine its degradation rate in both a skeletal and extraskeletal site. In experiment 1, eggshell particles with four different sizes (50, 75, 150, and 300 microns in diameter) were implanted in subcutaneous pouches of 30 rats. In experiment 2, a fragment of ostrich eggshell was implanted on the nasal dorsum of 10 rats. Animals were sacrificed at 1 (N = 10), 2 (N = 10), and 4 months (N = 10) during the first stage of the study, and at 1 year during the second stage of the study. The results were assessed by X-ray examination and routine histological techniques. In experiment 1, all animals healed uneventfully. At 1 month, only 50-micron particles had undergone resorption. At 2 months, both 50- and 75-micron particles had undergone resorption. At 4 months, the 150- and 300-micron particles were resorbed incompletely. Histologically, the eggshell elicited a mild inflammatory reaction at 1 month that decreased progressively at further stages. In experiment 2, all animals except one healed uneventfully. Radiologically, the eggshell implant displayed a noticeable stability. Histologically, seven of nine implants were encapsulated, but two of them were surrounded by a bony rim. In conclusion, eggshell is a resorbable implant, but the degradation kinetic is size dependent. Large ostrich grafts are also suitable as onlay graft, but a complementary osteosynthesis is recommended to enhance osteointegration.

The purpose of the study was to determine the efficacy of pentosan polysulphate, used in combination with guided bone regeneration on rat skull defects. The study was conducted on 45 adult Wistar rats. On each animal two symmetrical 6 mm wide, full-thickness, skull defects were created in the parietal regions. The right defect was chosen as the experimental site and the left one was left empty to provide a control. Each experimental site was covered by an inner and outer polytetrafluoroethylene membrane. The 45 rats were divided into 3 groups: in group I (n = 15), carboxymethyl cellulose, used as a delivery vehicle, was injected between the two membranes; in group II (n = 15), 1 mg of pentosan polysulfate was added to the carboxymethyl cellulose vehicle; in group III (n = 15), purified micronized eggshell powder was added to the mixture of pentosan polysulfate and carboxymethyl cellulose between the two membranes. In each group, the animals were sacrificed at 42 days. The harvested specimens were processed for contact radiography and standard histological examination. The results were assessed by a Fisher's exact test. All animals, except one, healed uneventfully. In group I, partial bone healing was observed in 14 out of 15 animals. In group II, partial bone healing was observed in 13 out of 15 animals, and complete bone healing in 1 out of 15 cases. In group III, partial resorption of the eggshell implant was observed with a partial bone healing in only 2 cases (P < 0.001). In conclusion, significant bone regeneration was observed with the membranes alone. The use of pentosane polysulphate did not result in additional bone gain. The use of particulate material as a space maintainer is also questionable.

Lymphoscintigraphy, after arousing great hope in the past in the field of breast cancer, has now been abandoned. The inability of this examination to predict the metastatic status of the nodes, and progress in therapeutic concepts have led to abandoning this technique. However, certain problems encountered by regional irradiation programmes and the work concerning sentinel node detection may bring this technique back into the spotlight. Lymphoscintigraphy may make it possible to adopt an individual approach, case by case, of the lymphatic drainage basins in breast tumors, thus enabling certain patients to benefit from regional irradiation when it would not have been traditionally recommended for this irradiation. Another aspect concerns the problem of the volumes irradiated. Work carried out with lymphoscintigraphy has enabled internal mammary chain nodes to be precisely located. Theses studies show the necessity of adapting the irradiation field to each individual case, but the clinical impact is limited, in the end, by the low recurrence rate in the internal mammary chain area. However, the new techniques of computer merging of scintigraphic and scanner images could enable the spatial position of the nodes in the upper axillary and supraclavicular regions to be determined. This would have, a priori, much wider clinical impact. Lymphoscintigraphic detection of the sentinel node is another field of major interest, but this technique is in competition with staining techniques. This procedure leads to a large reduction in morbidity of axillary surgery in 70% of patients. The use of techniques for detecting micrometastases in the sentinel node opens prospects in terms of prognosis. The qualities of differents radiotracers and different injection sites possible are also discussed.