Guidelines for May-Grünwald-Giemsa staining in haematology and non-gynaecological cytopathology: recommendations of the French Society of Clinical Cytology (SFCC) and of the French Association for Quality Assurance in Anatomic and Cytologic Pathology (AFAQAP).

OBJECTIVE: Since the guidelines of the International Committee for Standardisation in Haematology (ICSH) in 1984 and those of the European Committee for External Quality Assessment Programmes in Laboratory Medicine (EQALM) in 2004, no leading organisation has published technical recommendations for the preparation of air-dried cytological specimens using May-Grünwald-Giemsa (MGG) staining. DATA SOURCES: Literature data were retrieved using reference books, baseline-published studies, articles extracted from PubMed/Medline and Google Scholar, and online-available industry datasheets. RATIONALE: The present review addresses all pre-analytical issues concerning the use of Romanowsky's stains (including MGG) in haematology and non-gynaecological cytopathology. It aims at serving as actualised, best practice recommendations for the proper handling of air-dried cytological specimens. It, therefore, appears complementary to the staining criteria of the non-gynaecological diagnostic cytology handbook edited by the United Kingdom National External Quality Assessment Service (UK-NEQAS) in February 2015.

Survival of patients managed in France for duodenal neuroendocrine tumors (D-NET): a 20-year multicenter cohort study from the GTE group: a cohort study.

INTRODUCTION: Duodenal neuroendocrine tumours (D-NETs) have a low incidence; however, their diagnosis has been increasing. Features such as tumour location, size, type, histological grade, and stage were used to adapt the treatment to either endoscopic (ER) or surgical (SR) resections. There is no consensus regarding the definitive treatment. The authors' study aimed to describe the management of non-metastatic, well-differentiated D-NETs in France and its impact on patient survival. METHODS: A registry-based multicenter study using prospectively collected data between 2000 and 2019, including all patients managed for non-metastatic G1 and G2 D-NETs, was conducted in the GTE group. RESULTS: A total of 153 patients were included. Fifty-eight benefited from an ER, and 95 had an SR. No difference in recurrence-free survival (RFS) was observed regardless of treatment type. There was no significant difference between the two groups (ER vs. SR) in terms of location, size, grade, or lymphadenopathy, regardless of the type of incomplete resection performed or regarding the pre-therapeutic assessment of lymph node invasion in imaging. The surgery allowed for significantly more complete resection (patients with R1 resection in the SR group: 9 vs. 14 in the ER group, P <0.001). Among the 51 patients with positive lymph node dissection after SR, tumour size was less than or equal to 1 cm in 25 cases. Surgical complications were more numerous ( P =0.001). In the sub-group analysis of G1-G2 D-NETs between 11 and 19 mm, there was no significant difference in grade ( P =0.977) and location ( P =0.617) between the two groups (ER vs. SR). No significant difference was found in both morphological and functional imaging, focusing on the pre-therapeutic assessment of lymph node invasion ( P =0.387). CONCLUSION: Regardless of the resection type (ER or SR) of G1-G2 non-metastatic D-NETs, as well as the type of management of incomplete resection, which was greater in the ER group, long-term survival results were similar between ER and SR. Organ preservation seems to be the best choice owing to the slow evolution of these tumours.

Structural enhancement and video endoscopy: results of a large prospective comparative study.

INTRODUCTION: The structural-enhancement (SE) function electronically improves the video-endoscopic signal of Olympus processors (EXERA CV-160 or greater), enabling an increase in relief that may help in the detection of flat or ulcerated and raised lesions, especially those of small size. We assessed the diagnostic impact of this technique in the screening of lesions during basic video colonoscopy. METHODS: Maximum-level SE was programmed into processors on alternate weeks, and endoscopy dates were planned by an assistant unaware of the SE schedule, thus ensuring randomization. The endoscopists-senior practitioners with 3-29 years of digestive endoscopy practice-were informed of the experiment >3 weeks before it began and were not told about it again either before or during the study. This was to ensure that endoscopy examinations were performed without over-awareness of the technical conditions. GIF-100 to -160 Olympus endoscopes were used. RESULTS: During the study, 606 patients underwent upper digestive video-endoscopy, 305 with and 301 without the use of the SE function. Of 645 patients who underwent video colonoscopy, 593 were included in the study and 52 were excluded due to poor cleansing (8%); of those included, 330 were analyzed with and 263 without the SE function. We observed no differences in the detection of lesions (small or large) by either upper digestive endoscopy or video colonoscopy. CONCLUSION: This is the first study comparing video-endoscopy diagnosis with or without SE during upper digestive endoscopy and colonoscopy. The SE function available on Olympus video-endoscopy processors had no impact on the detection of lesions, not even on those of very small size.

All-trans retinoic acid targets gastric cancer stem cells and inhibits patient-derived gastric carcinoma tumor growth.

Gastric carcinoma is the third leading cause of cancer-related death worldwide. This cancer, most of the time metastatic, is essentially treated by surgery associated with conventional chemotherapy, and has a poor prognosis. The existence of cancer stem cells (CSC) expressing CD44 and a high aldehyde dehydrogenase (ALDH) activity has recently been demonstrated in gastric carcinoma and has opened new perspectives to develop targeted therapy. In this study, we evaluated the effects of all-trans-retinoic acid (ATRA) on CSCs in human gastric carcinoma. ATRA effects were evaluated on the proliferation and tumorigenic properties of gastric carcinoma cells from patient-derived tumors and cell lines in conventional 2D cultures, in 3D culture systems (tumorsphere assay) and in mouse xenograft models. ATRA inhibited both tumorspheres initiation and growth in vitro, which was associated with a cell-cycle arrest through the upregulation of cyclin-dependent kinase (CDK) inhibitors and the downregulation of cell-cycle progression activators. More importantly, ATRA downregulated the expression of the CSC markers CD44 and ALDH as well as stemness genes such as Klf4 and Sox2 and induced differentiation of tumorspheres. Finally, 2 weeks of daily ATRA treatment were sufficient to inhibit gastric tumor progression in vivo, which was associated with a decrease in CD44, ALDH1, Ki67 and PCNA expression in the remaining tumor cells. Administration of ATRA appears to be a potent strategy to efficiently inhibit tumor growth and more importantly to target gastric CSCs in both intestinal and diffuse types of gastric carcinoma.

Low prevalence of monoclonal B cells in Helicobacter pylori gastritis patients with duodenal ulcer.

We have studied the prevalence of B-cell clonality among a large group of 320 patients with Helicobacter pylori gastritis and duodenal ulcer. These patients underwent endoscopic examination with multiple gastric biopsies at diagnosis and were followed 2 and 12 months after therapy. Histopathologic examination of 809 sets of biopsy specimens showed lymphoid gastritis with lymphoid aggregates or follicles, but without lymphoepithelial lesion, in 302 samples corresponding to initial biopsy specimens (n=130) or to posttreatment biopsy specimens (n=172). DNA extracted from fresh antral specimens allowed the amplification of Helicobacter pylori DNA in all cases before therapy. The arrangement of the immunoglobulin heavy chain gene was studied by polymerase chain reaction (PCR) in the 302 selected lymphoid gastritis samples. Single or dominant bands were seen only in four specimens from three patients (1.3%), whereas a polyclonal pattern was seen in the other 298 samples. The detection threshold of our PCR technique was approximately 3% of clonal B cells diluted in a polyclonal population. This threshold appeared to be a reliable cutoff between polyclonal gastritis and clonal MALT lymphoma. In our experience, Helicobacter pylori lymphoid gastritis appeared mainly as a benign polyclonal condition.

Adenomatous hyperplasia in cirrhotic livers: histological evaluation, cellular density, and proliferative activity of 35 macronodular lesions in the cirrhotic explants of 10 adult French patients.

We examined 41 consecutive cirrhotic liver explants from French patients for the presence of nodules of adenomatous hyperplasia (AH) and then analyzed these lesions, together with underlying cirrhosis (C) and associated hepatocellular carcinoma (HCC), for various histological parameters, cellular density, and proliferative activity. Thirty-five AHs were identified in 10 livers (prevalence, 24%); seven of 10 were HCV positive. Hepatocellular carcinoma was more frequent in patients with AH than in patients without. The AHs consisted of 17 ordinary (OAH) and 18 atypical (AAH) adenomatous hyperplasia lesions. There was a malignant focus in five of the 18 AAHs. Wide areas of large liver cell dysplasia were frequent in OAH but never found in AAH. Obvious steatosis was frequent in HCC but exceptional in AAH and absent in OAH. There was a significant increase in cellular density in AAH and HCC as compared with C and OAH. Proliferative cell nuclear antigen immunostaining similarly showed an increase in proliferation from OAH or C to AAH and HCC. These data suggest that, in Europe as in Japan, one pathway of hepatocarcinogenesis is a multistep process in which AAH should be considered as a premalignant lesion very close to grade I HCC, while OAH seems to correspond to a regenerative nodule with limited proliferative ability.

CD30-positive cutaneous large cell lymphomas. A comparative study of clinicopathologic and molecular features of 16 cases.

The authors have analyzed and compared the clinicopathologic and molecular features of 16 cases of large cell cutaneous lymphomas expressing CD30 antigen. Three main clinical groups were defined: (1) a group of localized skin disease (7 cases); (2) a group of multicentric skin disease (5 cases); and (3) a group of concomitant skin and extracutaneous disease. Good prognosis was associated with localized skin disease and no history of lymphoma. Interestingly, a majority of Reed Sternberg-like cells was only observed in this group (5 of 6 cases). The two other groups did not show distinctive evolutive nor morphologic features. Southern blot and/or polymerase chain reaction (PCR) technique showed clonality and a T-cell genotype in respectively 13 of 14 and 12 of 12 analyzed cases. Viral infection of tumoral cells was investigated by PCR, in situ hybridization (ISH) or electron microscopy. Epstein-Barr virus (EBV) sequences were detected by PCR and ISH in tumoral cells of cutaneous lesions in one case of skin lymphoma with extracutaneous spreading. No EBV sequence was detected by ISH in the localized lymphomas, whereas HIV particles were visible in tumoral cells in one of these cases. No human T-cell lymphotropic virus (HTLV) tax sequence was amplified by PCR in any case of our series. Our results confirm that CD30-positive cutaneous large cell lymphomas are different clinical and molecular entities. However, a combined clinical and morphologic analysis may help to identify a subset of CD30 cutaneous lymphomas with favorable prognosis.

Axillary lymphadenectomy prepared by fat and lymph node suction in breast cancer.

AIMS: To describe a new technique of axillary dissection and evaluate the results. METHODS: Axillary lymphadenectomy prepared by fat and lymph node suction was performed on 43 patients with breast cancer and uninvolved axilla on physical examination. The mean follow-up was 18 months (range 12-27). After lipolysis and liposuction of the axilla, lymph node dissection was performed by axilloscopy in patients treated with conservative surgery. For patients undergoing modified radical mastectomy, a mastectomy with open axillary dissection was carried out. The axillary space was drained until less than 20 ml of lymphorrhea was being produced per day. Seventeen modified radical mastectomies (group M) and 26 lumpectomies (group L) were performed. RESULTS: No injury of muscles, vessels or nerves was observed. An average of 13.6 lymph nodes (8-31) were removed. In group L, the mean number of lymph nodes removed by liposuction, axilloscopy and control incision was 2.5, 5.4 and 4.2, respectively. In group M, the mean number of lymph nodes removed was 14. None of the 589 lymph nodes examined showed any pathological trauma. Fourteen patients (32%) had more than one histologically positive node. The mean quantity of lymphorrhea was 390 ml in group M and 275 ml in group L. The mean duration of drainage was 6 days in group M and 5 days in group L. Four patients had seromas which required punctures. Two patients had shoulder restriction (movements < 90 degrees) at 12 months. No arm oedema was seen. CONCLUSIONS: Axillary lymphadenectomy prepared by fat and lymph node suction is a reliable and effective procedure. However, it does not appear to be better than standard dissection as regards post-operative complications, except for arm oedema, but this must be confirmed by further studies.

Cytologic findings in a neovagina created with Vecchietti's technique for treating vaginal aplasia.

OBJECTIVE: To report long-term cytologic findings after treatment of congenital vaginal aplasia or Mayer-Rokitansky-Küster-Hauser syndrome (MRKH) using progressive traction of the vulvar tissues, or Vecchietti's technique (VT). STUDY DESIGN: In eight women with MRKH, neovaginal smears stained by the Harris-Schorr technique were collected 2-12 years after VT. RESULTS: Eosinophilic superficial cells observed in all cases indicate that the neovaginal epithelium responds to hormonal variations. The presence of Döderlein bacilli (seven cases) suggests that the regional environment of the neovagina is almost the same as that of the normal vagina. Gardnerella vaginalis (one case) and human papilloma virus infection (low grade squamous intraepithelial lesions) (one case) were also observed. CONCLUSION: Neovaginal smears taken after the cicatrization period following construction of a neovagina by VT show a remarkable resemblance to those from normal vaginal mucosa and suggest that these neovaginas are almost the same as normal vaginas.

[Clear cell adenoma of the parathyroid gland: a rare and misleading lesion]. / Adénome parathyroïdien à cellules claires: une lésion rare et trompeuse.

Parathyroid adenomas are common lesions and are considered to be the cause of most of the primary hyperparathyroidism cases. We report the case of a 73 year-old man who presented with a primary hyperparathyroidism. Clinical and histological explorations revealed the presence of an isolated parathyroid tumor containing exclusively clear cells and devoid of malignancy. This is the second reported case of clear cell parathyroid adenoma. Thus, in spite of its low occurrence, this diagnosis must be considered after rejection of the most frequent parathyroid clear cell hyperplasia and parathyroid carcinoma, or depending of the location, clear cell thyroid tumor and clear cell renal carcinoma metastasis.

[Study of B-lymphocyte clonality using in vitro gene amplification (PCR) in paraffin embedded samples]. / Etude de la clonalité B lymphocytaire par la technique d'amplification de gène in vitro (PCR) à partir de prélèvements fixés et inclus en paraffine.

Family-specific primers were used in polymerase chain reaction (PCR) to analyse clonality of immunoglobulin heavy chain (IgH) gene rearrangement. DNA templates were extracted from formalin-fixed paraffin-embedded biopsies from B-cell lymphomas (n = 19), T-cell lymphomas (n = 3), reactive lymphadenopathies (n = 20) and negative controls (n = 2). PCR was also performed on DNA extracted from fragments of the same biopsies that were either fixed in Bouin's solution (n = 24) or snap-frozen (n = 22). The latter were also studied for IgH gene rearrangement by the Southern blot technique. Polyclonal or clonal fragments were obtained from all frozen biopsies. No amplification was observed in 3 out of 44 formalin fixed specimens and 18 out of 24 Bouin fixed specimens. A clonal amplification was only observed in 16 out of 19 formalin-fixed B-cell lymphoma specimens. Therefore this technique allows to study B-cell clonality from formalin-fixed, embedded material.

[Transthoracic aspiration. Evaluation of cytologic and histologic diagnosis in a pulmonary nodule by retrospective comparison of 2 series of 267 cytoaspirations and 292 coaxial needle aspirations]. / La ponction trans-thoracique. Evaluation du diagnostic cytologique et histologique d'un nodule pulmonaire par comparaison rétrospective de 2 séries de 267 cytoponctions et de 292 ponctions à l'aiguille coaxiale.

Percutaneous fine-needle aspiration is a well established method for the diagnosis of peripheral lung lesion. In order to compare different methods of aspiration, we analyze retrospectively two different series: 267 fine needle aspirations (FNA) compared with the histological diagnosis on surgical specimens and 292 lung biopsies using a coaxial technique with comparison between cytological diagnosis--smears and imprints--and histological diagnosis simultaneously obtained on the same specimen. The sensitivity (91%), the specificity (90%) and the overall typing accuracy related to the histological types obtained by FNAB are equivalent to those of the literature. The low rate of pneumothorax in the series (6%) is related to the use of immediate interpretation of the specimen. Automated biopsy with a coaxial cutting needle provide cytological specimens--smears and imprint--with a high rate of sensibility (95.3%) and of sensibility (98%). The overall sensitivity of the cytological methods alone is better than biopsy (95.3% vs. 92.9%), but the typing accuracy is not as good as biopsy alone (98% vs. 100%). False-positive and false-negative diagnoses are the same in both series. In conclusion the percutaneous aspiration method choose to establish a morphological diagnosis in lung lesion depends now on the habits of the radiologist and the pathologist.

[Conservative treatment of adenocarcinomas of the anorectal junction by preoperative radiotherapy and intersphincteral resection]. / Traitement conservateur des adénocarcinomes de la jonction anorectale par radiothérapie préopératoire et résection intersphinctérienne.

AIM: Adenocarcinomas of the anorectal junction, especially T3 lesions, are usually treated by abdominoperineal resection. The aim of this study was to evaluate oncologic and functional results following conservative radiosurgical treatment of cancers of the anorectal junction. METHODS: From 1990 to 1999, among 395 patients with rectal carcinoma, 31 had sphincter-saving resection for a tumour located between 2 to 4.5 cm (mean 3.6) from the anal verge. There were 16 men and 15 women, mean age 62 years (range 30-86). There were 5 T2, 23 T3 and 3 T4 tumours; 17 were N1 and 3 were M1. Preoperative radiotherapy was performed in 26 patients (dose: 46 Gy, range: 36-54), with concomitant chemotherapy in 14 cases. Intersphincteric resection was performed six weeks after neoadjuvant treatment. Coloanal anastomoses were associated with a colonic pouch in 22 cases and with a protecting stoma in all cases. RESULTS: There was no postoperative mortality. Seven complications occurred: 3 anastomotic fistulas, 3 pelvic haemorrhages and 1 acute pancreatitis. Three patients had a definitive stoma. After preoperative radiotherapy, down-staging (pT0-2 N0) occurred in 46% of cases (12/26). Distal margin was 2.2 cm (range: 1-3) and was microscopically safe in all cases. Lateral margin was safe (> or = 1 mm) in 97% of cases. With a mean follow-up of 36 months, no local recurrence was suspected. Twenty-six patients (84%) were alive, 23 free of disease. Half of the patients had perfect continence, whereas the other half had occasional minor soiling. Functional results were better in patients with a colonic pouch. CONCLUSION: Conservative treatment of carcinomas of the anorectal junction is possible without compromising pelvic control and patient survival. Pelvic control was probably achieved by using preoperative radiotherapy with intersphincteric resection, ensuring safe distal and lateral margins.

[Lymphomatoid papulosis in a child]. / Papulose lymphomatoïde chez un enfant.

BACKGROUND: Lymphomatoid papulosis (LyP), uncommon in children, has a benign clinical course in contrast with a malignant histology. CASE REPORT: A 9-year old boy developed nodular skin lesions for 6 months on the scalp and penis and a papule skin on his trunk. Surgical excision of scalp nodules was performed to prevent necrosis and ulceration of the ear. The biopsy showed a dense mixed cellular infiltrate including histiocytes, lymphocytes and numerous atypical large cells of which a minority expressed the CD30 antigen. PCR showed an oligoclonal rearrangement of the TCR gamma chain gene. Physical examination, bone marrow aspirate and thoracoabdominal CT scans were normal. Three years after surgery, no other lesions occurred. DISCUSSION: Differential diagnosis of LyP is difficult with non Hodgkin's lymphoma, especially CD30+ lymphoma. Prognosis remains the major problem in LyP. Ten to 20% of cases are associated with lymphomas in adults.

[Does axillary liposuction modify histologic study of excised lymph nodes?]. / La lipoaspiration axillaire modifie-t-elle l'étude histologique des ganglions du curage?

OBJECTIVE: To determine the pathological features of lymph nodes removed by axillary liposuction. METHODS: A prospective study of 34 axillary dissections performed from July 1995 to September 1996 in patients with breast cancer N0. After lipolysis, the fat was drained from the axillary cavity by means of liposuction (Karman nozzle ch.8; suction pressure of 1 Bar). The remaining nodes were removed by an endoscopic dissection in conservative treatments. The remaining nodes were removed during modified radical mastectomy in non-conservative treatments. Lymph nodes were fixed in formol and examined by the pathologist. RESULTS: An average of 15 lymph nodes (8-35) were removed. 502 lymph nodes were examined: 458 (91%) were not involved and 44 (9%) were involved, including 21 (4%) with rupture of the capsule. No pathological traumatism was seen. CONCLUSION: Axillary liposuction did not alter the pathological features of lymph nodes in our study.

[Somatostatinoma. Apropos of a case]. / Le somatostatinome. A propos d'un cas.

Somatostatinoma are uncommon pancreatic endocrine tumors. We review the epidemiological, pathological, clinical and biological characteristics. The diagnosis of a somatostatinoma may be suggested clinically but is confirmed by histology of the resected tumor and specific immunohistochemistry marking. Malignancy is diagnosed on the presence metastases. Surgery is required with excision of the tumor, lymph nodes and metastases.

[A case of extraskeletal Ewing's sarcoma of the thoracic wall]. / A proposito di un caso di sarcoma extraosseo di Ewing della parete toracica.

The Authors report a case of extraskeletal Ewing's sarcoma of thoracic wall in a 15-year-old sport-man, who complained about pain in the left upper extremity. The diagnosis was performed by radiology (ultrasound study, TC, RM) and pre-operative needle-aspiration biopsy. The patient was treated by neoadjuvant chemotherapy, wide surgical resection and, then, radiotherapy. Neither local recurrence nor metastases have developed for 8 months.

Helicobacter pylori generates cells with cancer stem cell properties via epithelial-mesenchymal transition-like changes.

Helicobacter pylori infection is the major risk factor for gastric adenocarcinoma. The link with gastric adenocarcinoma is partly due to the H. pylori CagA oncoprotein. CagA is responsible for a particular cell phenotype in vitro, the 'hummingbird' phenotype, that corresponds to an elongation of the cells, mimicking an epithelial-mesenchymal transition (EMT). EMT participates in the carcinogenesis process, and is involved in the generation of cancer stem cells (CSCs). However, its involvement in gastric carcinogenesis has yet not been studied. Therefore, the aim of this study was to determine the role of H. pylori in EMT and in the emergence of gastric CSCs. For this purpose, gastric epithelial cells were cocultured with a cagA-positive H. pylori strain or its isogenic-deleted mutants or were transfected with CagA expression vectors. Study of the expression of epithelial and mesenchymal markers showed that H. pylori, via CagA, is responsible for an EMT phenotype associated with an increase in mesenchymal markers as well as CD44 expression, a known gastric CSC marker. Moreover, infection led to an increased ability to migrate, to invade and to form tumorspheres. Cell sorting experiments showed that only the CD44(high) cells induced by H. pylori infection displayed the mesenchymal phenotype and CSC properties in vitro, and had higher tumorigenic properties than CD44(low) cells in xenografted mice. Immunohistochemistry analyses on human and mouse gastric mucosa tissue samples confirmed a high expression of CD44 and mesenchymal markers in H. pylori-infected cases, and in gastric dysplasia and carcinoma. All of these data suggest that H. pylori, via CagA, unveils CSC-like properties by induction of EMT-like changes in gastric epithelial cells.

Mucosal healing with methotrexate in Crohn's disease: a prospective comparative study with azathioprine and infliximab.

BACKGROUND: Mucosal healing has become a new therapeutic goal in Crohn's disease and can be achieved with azathioprine (AZA) or biologics. Methotrexate (MTX) is an effective drug for both the induction and maintenance of remission in Crohn's disease. However, mucosal healing with MTX has been poorly investigated. AIM: To assess the mucosal healing rate in patients with Crohn's disease with clinical response to MTX as compared with AZA or infliximab (IFX). METHODS: From October 2007 to May 2009, consecutive patients with Crohn's disease were prospectively enrolled into a single-centre study when they met the following criteria: previous identification of mucosal ulcerations with ileo-colonoscopy, clinical remission within at least 3 months with MTX, AZA or IFX monotherapy, usual indication for colonoscopy in Crohn's disease (dysplasia/cancer screening, suspected stenosis) excluding assessment for mucosal healing. Mucosal healing was defined as absence of mucosal ulceration in all segments. RESULTS: Fifty-one patients with Crohn's disease (38 female; median age: 42 years) were included: 18 receiving MTX, 18 AZA and 15 IFX. Mucosal healing was achieved in 2/18 (11%) with MTX, in 9/18 (50%) with AZA (P =0.011 vs. MTX) and in 9/15 (60%) with IFX (P=0.008 vs. MTX). CONCLUSION: In patients with Crohn's disease in sustained clinical remission, mucosal healing is less frequently achieved with MTX as compared with AZA or IFX.