Extracellular vesicles derived from injured vascular tissue promote the formation of tertiary lymphoid structures in vascular allografts.

Tertiary lymphoid structures (TLS) accumulate at sites of chronic injury where they function as an ectopic germinal center, fostering local autoimmune responses. Vascular injury leads to the release of endothelial-derived apoptotic exosome-like vesicles (ApoExo) that contribute to rejection in transplanted organs. The purpose of the study was to evaluate the impact of ApoExo on TLS formation in a model of vascular allograft rejection. Mice transplanted with an allogeneic aortic transplant were injected with ApoExo. The formation of TLS was significantly increased by ApoExo injection along with vascular remodeling and increased levels of antinuclear antibodies and anti-perlecan/LG3 autoantibodies. ApoExo also enhanced allograft infiltration by γδT17 cells. Recipients deficient in γδT cells showed reduced TLS formation and lower autoantibodies levels following ApoExo injection. ApoExo are characterized by proteasome activity, which can be blocked by bortezomib. Bortezomib treated ApoExo reduced the recruitment of γδT17 cells to the allograft, lowered TLS formation, and reduced autoantibody production. This study identifies vascular injury-derived extracellular vesicles (ApoExo), as initiators of TLS formation and demonstrates the pivotal role of γδT17 in coordinating TLS formation and autoantibody production. Finally, our results suggest proteasome inhibition with bortezomib as a potential option for controlling TLS formation in rejected allografts.

MCM2: An alternative to Ki-67 for measuring breast cancer cell proliferation.

Breast cancer is a heterogeneous disease comprising a diversity of tumor subtypes that manifest themselves in a wide variety of clinical, pathological, and molecular features. One important subset, luminal breast cancers, comprises two clinically distinct subtypes luminal A and B each of them endowed with its own genetic program of differentiation and proliferation. Luminal breast cancers were operationally defined as follows: Luminal A: ER+, PR+, HER2-, Ki-67<14% and Luminal B: ER+ and/or PR+, HER2-,Ki-67≥14% or, alternatively ER+ and/or PR+, HER2+, any Ki-67. There is currently a need for a clinically robust and validated immunohistochemical assay that can help distinguish between luminal A and B breast cancer. MCM2 is a family member of the minichromosome maintenance protein complex whose role in DNA replication and cell proliferation is firmly established. As MCM2 appears to be an attractive alternative to Ki-67, we sought to study the expression of MCM2 and Ki-67 in different histological grades and molecular subtypes of breast cancer focusing primarily on ER-positive tumors. MCM2 and Ki-67 mRNA expression were studied using in silico analysis of available DNA microarray and RNA-sequencing data of human breast cancer. We next used immunohistochemistry to evaluate protein expression of MCM2 and Ki-67 on tissue microarrays of invasive breast carcinoma. We found that MCM2 and Ki-67 are highly expressed in breast tumors of high histological grades, comprising clinically aggressive tumors such as triple-negative, HER2-positive and luminal B subtypes. MCM2 expression was detected at higher levels than that of Ki-67 in normal breast tissues and in breast cancers. The bimodal distribution of MCM2 scores in ER+/HER2- breast tumors led to the identification of two distinct subgroups with different relapse-free survival rates. In conclusion, MCM2 expression can help sorting out two clinically important subsets of luminal breast cancer whose treatment and clinical outcomes are likely to diverge.

MMP-9 expression varies according to molecular subtypes of breast cancer.

BACKGROUND: In 2014, breast cancer remains a major cause of mortality worldwide mostly due to tumor relapse and metastasis. There is currently a great interest in identifying cancer biomarkers and signalling pathways mechanistically related to breast cancer progression. Matrix metalloproteinase-9 (MMP-9) is a member of matrix degrading enzymes involved in cancer development, invasion and metastasis. Our objective was to investigate MMP-9 expression in normal human breast tissue and to compare it to that of breast cancer of various histological grades and molecular subtypes. We also sought to correlate MMP-9 expression with the incidence of metastasis, survival rates and relapse in breast cancer patients. METHODS: MMP-9 was first studied using in silico analysis on available DNA microarray and RNA sequencing data of human breast cancer tissues and human breast cancer cell lines. We next ascertained MMP-9 expression in both normal breast tissue and in human breast carcinoma tissue microarrays. RESULTS: Significant increase in MMP-9 expression was found in breast cancer cells where compared to normal breast tissue. A positive correlation could also be established between elevated levels of MMP-9 and breast cancer of high histological grade. Furthermore, our results indicate that not only MMP-9 is differentially expressed between each molecular subset but also, more importantly MMP-9 overexpression revealed itself as a startling feature of triple-negative and HER2-positive breast cancers. Lastly, the clinical relevance of MMP-9 overexpression is strongly supported by its significant association with a higher incidence of metastasis and relapse. CONCLUSIONS: Differential expression of MMP-9 reflects the extent of cellular differentiation in breast cancer cells and is closely related to the most aggressive subtypes of breast cancer. Hence, MMP-9 is a promising prognostic biomarker of high-grade breast cancer. In our opinion, MMP-9 expression could help segregate subsets of aggressive breast cancer into clinically meaningful subtypes.

eIF4E phosphorylation promotes tumorigenesis and is associated with prostate cancer progression.

Translational regulation plays a critical role in the control of cell growth and proliferation. A key player in translational control is eIF4E, the mRNA 5' cap-binding protein. Aberrant expression of eIF4E promotes tumorigenesis and has been implicated in cancer development and progression. The activity of eIF4E is dysregulated in cancer. Regulation of eIF4E is partly achieved through phosphorylation. However, the physiological significance of eIF4E phosphorylation in mammals is not clear. Here, we show that knock-in mice expressing a nonphosphorylatable form of eIF4E are resistant to tumorigenesis in a prostate cancer model. By using a genome-wide analysis of translated mRNAs, we show that the phosphorylation of eIF4E is required for translational up-regulation of several proteins implicated in tumorigenesis. Accordingly, increased phospho-eIF4E levels correlate with disease progression in patients with prostate cancer. Our findings establish eIF4E phosphorylation as a critical event in tumorigenesis. These findings raise the possibility that chemical compounds that prevent the phosphorylation of eIF4E could act as anticancer drugs.

MCM2, MCM4, and MCM6 in Breast Cancer: Clinical Utility in Diagnosis and Prognosis.

Breast cancer is a heterogeneous disease comprising the estrogen receptor (ER)-positive luminal subtype which is subdivided into luminal A and luminal B and ER-negative breast cancer which includes the triple-negative subtype. This study has four aims: 1) to examine whether Minichromosome Maintenance (MCM)2, MCM4, and MCM6 can be used as markers to differentiate between luminal A and luminal B subtypes; 2) to study whether MCM2, MCM4, and MCM6 are highly expressed in triple-negative breast cancer, as there is an urgent need to search for surrogate markers in this aggressive subtype, for drug development purposes; 3) to compare the prognostic values of these markers in predicting relapse-free survival; and 4) to compare the three approaches used for scoring the protein expression of these markers by immunohistochemistry (IHC). MCM2, MCM4, MCM6, and MKI67 mRNA expression was first studied using in silico analysis of available breast cancer datasets. We next used IHC to evaluate their protein expression on tissue microarrays using three scoring methods. MCM2, MCM4, and MCM6 can help in distinction between luminal A and luminal B whose therapeutic management and clinical outcomes are different. MCM2, MCM4, MCM6, and Ki-67 are highly expressed in breast cancer of high histological grades that comprise clinically aggressive tumors such as luminal B, HER2-positive, and triple-negative subtypes. Low transcript expression of these markers is associated with increased probability of relapse-free survival. A positive relationship exists among the three scoring methods of each of the four markers. An independent validation cohort is needed to confirm their clinical utility.

The double capsules in macro-textured breast implants.

Breast implants are amongst the most widely used types of permanent implants in modern medicine and have both aesthetic and reconstructive applications with excellent biocompatibility. The double capsule is a complication associated with textured prostheses that leads to implant displacement; however, its etiology has yet to be elucidated. In this study, 10 double capsules were sampled from breast expander implants for in-depth analysis; histologically, the inner capsular layer demonstrated highly organized collagen in sheets with delamination of fibers. At the prosthesis interface (PI) where the implant shell contacts the inner capsular layer, scanning electron microscopy (SEM) revealed a thin layer which mirrored the three-dimensional characteristics of the implant texture; the external surface of the inner capsular layer facing the intercapsular space (ICS) was flat. SEM examination of the inner capsule layer revealed both a large bacterial presence as well as biofilm deposition at the PI; a significantly lower quantity of bacteria and biofilm were found at the ICS interface. These findings suggest that the double capsule phenomenon's etiopathogenesis is of mechanical origin. Delamination of the periprosthetic capsule leads to the creation of the ICS; the maintained separation of the 2 layers subsequently alters the biostability of the macro-textured breast implant.

Comprehensive BRCA1 and BRCA2 mutation analyses and review of French Canadian families with at least three cases of breast cancer.

Few studies have reported on the comprehensive BRCA1/2 mutation analyses of hereditary breast cancer (HBC) families of French Canadian descent. Here we report the investigation of 82 families with at least 3 cases of breast cancer evaluated for mutations by DNA sequencing and/or multiplex ligation-dependent probe amplification (MLPA) assay. DNA sequencing identified pathogenic mutations in 37 (45.1%) families, of which 70.2% were one of three recurring mutations (BRCA1:R1443X, BRCA2:8765delAG, and BRCA2:E1953X) frequently reported in this founder population; and variants of uncertain clinical significance in 7 (8.5%) families of which two harbored BRCA2:E3002K. MLPA analysis of the 38 DNA sequence-negative families did not reveal any large rearrangements in BRCA1/2. A phenotypic characterization of the cancer families based on pathogenic mutation status revealed that there were significantly fewer very young age at diagnosis breast cancer cases (<36 years) in mutation-negative families (5.9%, 9 of 153) than in BRCA1 (22.8%, 13 of 57; P = 0.0003) or BRCA2 (22.9%, 27 of 118; P < 1× 10E5) mutation-positive families. There were significantly more mutation-positive families (29 of 36, 80.6%) with a very young age of onset of breast cancer case than those that did not (8 of 39, 20.5%) (P < 10E6). The comprehensive mutation analysis of BRCA1/2 suggests that genomic rearrangements are unlikely to account for sequence-negative HBC families and affirms that the presence of a very young age of diagnosis of breast cancer is strongly predictive of mutation carrier status of French Canadian HBC families.

Implant degradation and poor healing after endovascular repair of abdominal aortic aneurysms: an analysis of explanted stent-grafts.

PURPOSE: To study explanted stent-grafts to achieve a better understanding of the mechanisms of failure after endovascular treatment of abdominal aortic aneurysms (AAA). METHODS: Twelve stent-grafts were harvested at autopsy (n=3) or during surgical conversion (n=9). Device alterations were investigated by macroscopic examination, radiography, and surface analysis techniques. Healing around the implants was studied via histology and immunohistochemistry, with particular attention to the stent-graft/tissue interface. RESULTS: Degradation was more important with Vanguard stent-grafts (off the market) than with AneuRx and Talent stent-grafts, but rupture of nitinol wires and poor surface finish in Talent stent-grafts raise concern about their corrosion resistance and long-term stability. Poor healing was observed around stent-grafts even after several years of implantation, with absence of vascular smooth muscle cells, fibroblasts, and collagen formation. In addition to the well-known foreign body reaction around the graft, numerous polymorphonuclear cells characteristic of the first step of healing were present in tissues around stent-grafts retrieved at surgical conversion. Factors explaining the lack of tissue organization around stent-grafts are discussed. CONCLUSION: The long-term stability of implants remains a concern and requires more transparency from manufacturers regarding the surface properties of their devices. Lack of neointima formation impairs biological fixation of the implant to the vessel wall, leading to possible endoleaks and migration. New-generation stent-grafts promoting biological fixation should be developed to improve clinical outcomes of this minimally invasive treatment.

Basal cells of second trimester fetal breasts: immunohistochemical study of myoepithelial precursors.

The molecular characterization of human mammary myoepithelial cells is incomplete, hindering our understanding of its importance in breast physiology and pathology. Because data on the precursors of this cell lineage remain scarce and often contradictory, basal epithelial cells of second trimester fetal breasts were studied by light microscopy (LM) and immunohistochemistry (IHC). Up to 20 wk of gestational age, the mammary rudiments only comprised roundish primary outgrowths, "primary buds," more likely to represent immature nipples than true mammary tissue. At 21 wk secondary outgrowths, "projections," extended from enlarged primary buds into well-vascularized layers of dense mesenchyme. Basal projection cells had a partial myoepithelial-like phenotype: they reacted with CD29, CD49f, CD104, keratin 14, vimentin, S100beta protein, and p63; furthermore, many became positive for keratin 17, alpha-smooth muscle actin, and CD10 (but not for keratin 19) between wk 21 and 25. The continuous basement membrane associated with the fetal mammary rudiments was strongly positive for collagens type IV and VII, and for laminin 5. Consistently strong and basally polarized staining for hemidesmosomal components suggested that although incompletely differentiated, most second trimester myoepithelial precursors might already mediate local epithelial-mesenchymal interactions, i.e., complex signaling pathways which are crucial for both orderly growth during development and maintenance of homeostasis during adult life. Because they are likely implicated in the phenomenon of menstrual cycle-related growth spurts in the adult resting breast, the strategically positioned cells of the myoepithelial lineage might constitute critical protagonists in defective epithelial-mesenchymal signaling associated with cancer progression.