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In chronic lymphocytic leukaemia (CLL) the efficacy of SARS-CoV-2 vaccination remains unclear as most studies have focused on humoral responses. Here we comprehensively examined humoral and cellular responses to vaccine in CLL patients. Seroconversion was observed in 55.2% of CLL with lower rate and antibody titres in treated patients. T-cell responses were detected in a significant fraction of patients. CD4 + and CD8 + frequencies were significantly increased independent of serology with higher levels of CD4 + cells in patients under a Bruton tyrosine kinase (BTK) or a B-cell lymphoma 2 (BCL-2) inhibitor. Vaccination skewed CD8 + cells towards a highly cytotoxic phenotype, more pronounced in seroconverted patients. A high proportion of patients showed spike-specific CD4 + and CD8 + cells producing interferon gamma (IFNγ) and tumour necrosis factor alpha (TNFα). Patients under a BTK inhibitor showed increased production of IFNγ and TNFα by CD4 + cells. Vaccination induced a Th1 polarization reverting the Th2 CLL T-cell profile in the majority of patients with lower IL-4 production in untreated and BTK-inhibitor-treated patients. Such robust T-cell responses may have contributed to remarkable protection against hospitalization and death in a cohort of 540 patients. Combining T-cell metrics with seroprevalence may yield a more accurate measure of population immunity in CLL, providing consequential insights for public health., (© 2022 The Authors. British Journal of Haematology published by British Society for Haematology and John Wiley & Sons Ltd.)

Maximizing water use efficiency, yield, and plant survival under drought is a relevant research issue for almond-tree-growing areas worldwide. The intraspecific diversity of this species may constitute a valuable resource to address the resilience and productivity challenges that climate change poses to crop sustainability. A comparative evaluation of physiological and productive performance of four almond varieties: 'Arrubia', 'Cossu', 'Texas', and 'Tuono', field-grown in Sardinia, Italy, was performed. A great variability in the plasticity to cope with soil water scarcity and a diverse capacity to adapt to drought and heat stresses during fruit development were highlighted. The two Sardinian varieties, Arrubia and Cossu, showed differences in water stress tolerance, photosynthetic and photochemical activity, and crop yield. 'Arrubia' and 'Texas' showed greater physiological acclimation to water stress while maintaining higher yields, as compared to the self-fertile 'Tuono'. The important role of crop load and specific anatomical traits affecting leaf hydraulic conductance and leaf gas exchanges efficiency (i.e., dominant shoot type, leaf size and roughness) was evidenced. The study highlights the importance of characterizing the relationships among almond cultivar traits that affect plant performance under drought in order to better assist planting choices and orchard irrigation management for given environmental contexts.

Nucleophosmin (NPM1) mutations occurring in acute myeloid leukemia (AML) (about 50 so far identified) cluster almost exclusively in exon 12 and lead to common changes at the NPM1 mutants C-terminus, i.e., loss of tryptophans 288 and 290 (or 290 alone) and creation of a new nuclear export signal (NES), at the bases of exportin-1(XPO1)-mediated aberrant cytoplasmic NPM1 . Immunohistochemistry (IHC) detects cytoplasmic NPM1 and is predictive of the molecular alteration. Besides IHC and molecular sequencing, Western blotting (WB) with anti- NPM1 mutant specific antibodies is another approach to identify NPM1 -mutated AML. Here, we show that among 382 AML cases with NPM1 exon 12 mutations, one was not recognized by WB, and describe the discovery of a novel combination of two mutations involving exon 12. This appeared as a conventional mutation A with the known TCTG nucleotides insertion/duplication accompanied by a second event (i.e., an 8-nucleotide deletion occurring 15 nucleotides downstream of the TCTG insertion), resulting in a new C-terminal protein sequence. Strikingly, the sequence included a functional NES ensuring cytoplasmic relocation of the new mutant supporting the role of cytoplasmic NPM1 as critical in AML leukemogenesis.

NOTCH1 mutations and deregulated signal have been commonly found in chronic lymphocytic leukemia (CLL) patients. Whereas the impact of NOTCH1 mutations on clinical course of CLL has been widely studied, the prognostic role of NOTCH1 activation in CLL remains to be defined. Here, we analyzed the activation of NOTCH1/NOTCH2 (ICN1/ICN2) and the expression of JAGGED1 (JAG1) in 163 CLL patients and evaluated their impact on TTFT (Time To First Treatment) and OS (Overall Survival). NOTCH1 activation (ICN1+) was found in 120/163 (73.6%) patients. Among them, 63 (52.5%) were NOTCH1 mutated (ICN1+/mutated) and 57 (47.5%) were NOTCH1 wild type (ICN1+/WT). ICN1+ patients had a significant reduction of TTFT compared to ICN1-negative (ICN1-). In the absence of NOTCH1 mutations, we found that the ICN1+/WT group had a significantly reduced TTFT compared to ICN1- patients. The analysis of IGHV mutational status showed that the distribution of the mutated/unmutated IGHV pattern was similar in ICN1+/WT and ICN1- patients. Additionally, TTFT was significantly reduced in ICN1+/ICN2+ and ICN1+/JAG1+ patients compared to ICN1-/ICN2- and ICN1-/JAG1- groups. Our data revealed for the first time that NOTCH1 activation is a negative prognosticator in CLL and is not correlated to NOTCH1 and IGHV mutational status. Activation of NOTCH2 and JAGGED1 expression might also influence clinical outcomes in this group, indicating the need for further dedicated studies. The evaluation of different NOTCH network components might represent a new approach to refine CLL risk stratification., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Baldoni, Del Papa, De Falco, Dorillo, Sorrentino, Rompietti, Adamo, Nogarotto, Cecchini, Mondani, Silva Barcelos, Moretti, Mameli, Fabi, Sorcini, Stella, Giancola, Guardalupi, Ulbar, Plebani, Guarente, Rosati, Di Nicola, Marchioni, Di Ianni and Sportoletti.)

Context: Indoleamine 2,3-dioxygenase 1 (IDO1) is a single chain oxidoreductase that catalyzes tryptophan degradation to kynurenine. In cancer, it appears to exert an immunosuppressive function as part of an acquired mechanism of immune escape mediated by the inhibition of lymphocyte proliferation and survival and by the induction of FoxP3+ T regulatory cells., Objective: The objective of the study was to evaluate IDO1 expression in thyroid carcinoma and demonstrate its immunosuppressive function in the context of thyroid tumors., Setting: IDO1 expression was evaluated by quantitative PCR in 105 papillary thyroid carcinomas (PTCs), 11 medullary thyroid carcinomas, six anaplastic thyroid carcinomas, and five thyroid carcinoma cell lines (TCCLs), by immunohistochemistry in 55 PTCs and by Western blotting in five TCCLs. FoxP3+ Treg lymphocyte density was evaluated by immunohistochemistry in 29 PTCs. IDO1 inhibitory effect on lymphocyte proliferation was tested in coculture experiments of TCCLs and activated lymphocytes., Results: IDO1 mRNA expression resulted significantly higher in all the analyzed thyroid carcinoma histotypes compared with normal thyroid. Interestingly, an increase of IDO1 mRNA expression magnitude could be observed with gain of aggressiveness (PTCs and medullary thyroid carcinomas ≪ anaplastic thyroid carcinomas). In PTCs, IDO1 mRNA expression magnitude correlated with IDO1 immunostaining intensity in cancer cells and with FoxP3+ Treg lymphocyte density in the tumor microenvironment. IDO1 was expressed in human thyroid cancer cell lines in vitro, and FTC-133 cells showed high kynurenine concentration in the conditioned medium and a strong suppressive action on the proliferation of activated lymphocytes in coculture experiments., Conclusions: For the first time, this study demonstrates a pivotal role of IDO1 in the suppression of lymphocyte function in thyroid carcinoma microenvironment.

Correct function of spermatogonia is critical for the maintenance of spermatogenesis throughout life, but the cellular pathways regulating undifferentiated spermatogonia proliferation, differentiation, and survival are only partially known. We show here that long glucocorticoid-induced leucine zipper (L-GILZ) is highly expressed in spermatogonia and primary spermatocytes and controls spermatogenesis. Gilz deficiency in knock-out (gilz KO) mice leads to a complete loss of germ cell lineage within first cycles of spermatogenesis, resulting in male sterility. Spermatogenesis failure is intrinsic to germ cells and is associated with increased proliferation and aberrant differentiation of undifferentiated spermatogonia and with hyperactivity of Ras signaling pathway as indicated by an increase of ERK and Akt phosphorylation. Spermatogonia differentiation does not proceed beyond the prophase of the first meiotic division due to massive apoptosis associated with accumulation of unrepaired chromosomal damage. These results identify L-GILZ as a novel important factor for undifferentiated spermatogonia function and spermatogenesis.

Aims and Background: HER-3 signaling might contribute to resistance to trastuzumab. To clarify the role of HER-3 in HER-2-positive breast cancer, it is important to evaluate the level of HER-3 and its correlations with clinical outcome in metastatic breast cancer patients treated with trastuzumab., Methods: HER-3 status by immunohistochemistry was evaluated in HER-2-positive metastatic breast cancer patients treated with trastuzumab-based therapy at our institution. Two scorings were utilized for interpreting staining for HER-3, and the correlation between HER-3 status and clinical outcome was evaluated., Results: We evaluated HER-3 status in 61 of 76 HER-2-positive metastatic breast cancers treated with trastuzumab-based therapy at our institution from 4/1999 to 3/2006. We observed 55.2% objective responses; median time to progression and overall survival from start of trastuzumab therapy were 9.6 months (0.921-78.87) and 29.1 months (1.4-129.5+), respectively. With a cutoff of 50% staining tumor cells, we found 30 HER-3-negative and 31 HER-3-positive tumors. HER-3 status was not significantly associated with clinical outcome, but a shorter time to progression and overall survival were observed in patients with HER-3-positive tumors., Conclusions: HER-3 status by immunohistochemistry was not significantly associated with clinical outcome in HER-2-positive metastatic breast cancer patients. Further studies are necessary to evaluate the prognostic and predictive role of HER-3.

Epithelial cysts account for about 20% of all splenic cysts. Their pathogenesis is unclear, and different authors have proposed many hypotheses. It has been suggested that they are derived from embryonal epithelial inclusions during splenic development, from invagination of capsular surface mesothelium in splenic sulci with subsequent metaplasia, or from trauma. Moreover, a congenital, genetic, or teratomatous origin has also been hypothesized. We describe an unusual case of epithelial splenic cyst with mature liver foci in its wall. This finding supports its possible dysontogenetic origin., (Copyright © 2011 Elsevier Inc. All rights reserved.)

Purpose: The purpose of this study was to evaluate the preclinical safety of intravitreal genistein in rabbit eyes over a short-term period., Methods: Twelve New Zealand albino rabbits were selected for this study. Four concentrations of genistein (LC Laboratories, Woburn, MA) were prepared: 24 mg/0.1 mL, 135 mg/0.1 mL, 270 mg/0.1 mL, and 540 mg/0.1 mL. Each concentration was injected intravitreally in one eye of three rabbits. As a control, the vehicle solution was injected into the other eye of each animal. Retinal safety of intravitreal genistein was studied with electroretinography and histologic examination in rabbits. Electroretinography recordings were made before the injection and 3 weeks after the injection. Eventually, the rabbits were killed and the retinas were examined by light microscopy. Immunohistochemical staining with caspase-3 and caspase-9 was also performed to evaluate apoptotic expression in all study and control eyes., Results: Electroretinography studies showed no significant difference between control and genistein-injected eyes at any of the doses in the rabbit model. Histologic examination showed no retinal abnormality in the rabbits injected with different concentrations of genistein. Immunohistochemical staining with caspase-3 and caspase-9 showed no different apoptotic protein expression in any study or control eyes., Conclusion: Our results indicate that genistein is a safe intravitreal drug in the rabbit model up to 540 mg. If proven safe and efficacious in human studies, intravitreal injection of genistein could be considered a treatment alternative for ocular neovascularisation in selected cases.

Skeletal muscle undergoes regeneration generally after an injury and in some cases it may mimic a malignant process. We observed these aspects in association with abdominal wall endometriosis and as no similar conditions were found in the literature this prompted us to study the main clinicopathologic and immunohistochemical profile of this phenomenon. Thirteen cases of abdominal wall endometriosis were retrieved from the files of our Institute. All original slides were reviewed to reveal the presence of skeletal muscle and 8 cases were enrolled for morphologic and immunohistochemical studies as follows: vimentin, desmin, myoglobin, myogenin, myoD1, CD56, S100, and p21. Histologically, in 4 of the 8 cases in the skeletal muscle adjacent to the endometriotic foci there was a proliferation of round cells with the typical appearance of maturing myoblasts. More peripherally, myotubes and early myocytes were present. This proliferation was florid in 1 case and focal in 3 cases. At immunohistochemical investigation, the less differentiated cells reacted with vimentin, desmin, S100, CD56, myoD1, and myogenin but not with myoglobin or p21. On the contrary, intermediately differentiated cells showed a progressive loss of vimentin, CD56, and myoD1 whereas they were positive for desmin, S100, myogenin, myoglobin, and p21. Terminally differentiated cells reacted only with desmin and myoglobin. This peculiar immunohistochemical profile was consistent with the immunophenotype of maturing myoblasts, confirmed the regenerative nature of the phenomenon and allowed differential diagnosis with other proliferations sharing a similar morphology. The expression of early differentiation markers was greatest in the islands of cells nearest to endometriosis, whereas in the more distant areas the markers of late differentiation prevailed. This gradient of expression suggests that muscle cells are stimulated by growth factors or other signals produced by the cycling endometrioid foci. In conclusion, we report a hitherto undescribed entity that may mimic a malignant process, especially when the reaction is florid or when endometriotic glands and stroma are not clearly evident, as during the examination of small biopsies, frozen sections, or cytology samples. Therefore, although the histologic diagnosis of endometriosis is usually straightforward, pathologists should be aware of the concomitant regenerative effects on skeletal muscle, which may represent a possible diagnostic pitfall.

Objectives: The squamous cell carcinoma of the tongue (SCCT) is biologically and epidemiologically distinct from other oral cavity cancers and is associated with lower overall survival rates. The role of HER family members (HER-1, HER-2/neu, HER-3 and HER-4) in the pathogenesis and progression of head and neck squamous cell carcinomas has been demonstrated but no report have focused on SCCT. This study investigated, the expression of all members of the HER family, in a series of SCCT and studied the possible prognostic value and correlation with various clinico-pathological parameters., Methods: HER-1, HER-2/neu, HER-3 and HER-4 expression was analysed by semi-quantitative immunohistochemical staining on paraffin embedded tissue specimens from 40 patients who underwent surgery for SCCT between 1996 and 2006., Results: HER-1 was overexpressed in 26 cases (65%), HER-2/neu in two (5%), HER-3 in 19 (48%) and HER-4 in three cases (8%). No significant correlation was found between clinicopathological variables and expression of HER-1 and HER-2/neu. HER-3 overexpression was significantly related to nodal stage, age (>or=64 years) and decreased overall survival (P

Background: Insulin-like growth factor receptor-1 (IGFR-1) represents a novel molecular target in non-small-cell lung cancer (NSCLC). IGFR-1 and epidermal growth factor receptor (EGFR) activation is essential to mediate tumor cell survival, proliferation and invasion. We explored the correlation between IGFR-1 and EGFR, their relationship with clinicopathological parameters and their impact on outcome in resected stage I-III NSCLC patients., Patients and Methods: Tumors from 125 surgical NSCLC patients were evaluated for IGFR-1 and EGFR expression by immunohistochemistry. Kaplan-Meier estimates of survival and time to recurrence were calculated for clinical variables and biologic markers using the Cox model for multivariate analysis., Results: IGFR-1 protein overexpression was detected in 36.0% of NSCLC patients and was associated with larger tumor size (P = 0.04) but not with other clinical or biological characteristics. EGFR protein overexpression was observed in 55.2% of NSCLC, more frequently in squamous cell carcinoma (SCC) than non-SCC (63.7% versus 36.3%, chi(2) = 9.8, P = 0.001). IGFR-1 protein expression was associated with EGFR protein expression (P = 0.03). At the multivariate analysis, high coexpression of both IGFR-1 and EGFR was a significant prognostic factor of worse disease-free survival (DFS) (hazard ratio 2.51, P = 0.01)., Conclusion: A statistically significant association was observed between high coexpression of both IGFR-1 and EGFR and worse DFS in early NSCLC patients.

Background: In an attempt to identify markers of resistance to trastuzumab, we evaluated both the profiling of human epidermal growth factor receptor 2 (HER2)-positive tumor cells measuring the relative levels of EGFR, pMAPK, pAkt and PTEN and their correlations with clinical outcome in HER2-positive metastatic breast cancer patients treated with trastuzumab., Patients and Methods: Tumor tissues for this retrospective analysis were available from 45 out of 76 patients with metastatic breast cancer treated from April 1999 to March 2006 with trastuzumab-based therapy at our Institution. Evaluations of EGFR, pMAPK, pAkt and PTEN status by immunohistochemistry (IHC) were carried out on all 45 tissue samples and their correlations with response to trastuzumab, incidence of central nervous system (CNS) metastases, time to progression (TTP), overall survival from diagnosis of breast cancer (OS1), from diagnosis of metastatic disease (OS2) and from the start of trastuzumab (OS3) were analyzed., Results: We observed that TTP (P = 0.001) and median OS2 and OS3 were significantly longer in patients responsive to trastuzumab-based regimen compared with nonresponsive patients. EGFR, pMAPK, pAkt and PTEN status by IHC were not significantly associated with response to trastuzumab, TTP, overall survival (OS1, OS2, OS3) and CNS metastases incidence. A trend for shorter OS3 was observed for pMAPK-positive patients compared with pMAPK-negative patients (22.8 versus 31.2 months; P = 0.076). Median OS1 resulted shorter in 22 pAkt-positive patients (69.8 months) compared with 23 pAkt-negative patients (108.2 months); P = 0.091. It is likely that high expression of pMAPK (pMAPK-positive status) or pAkt (pAkt-positive status) could identify a subgroup of HER2-positive tumors with high activity of proliferation and survival pathways and with resistance to trastuzumab., Conclusions: In HER2-positive metastatic breast cancers, EGFR, pMAPK, pAkt and PTEN status evaluated by IHC was not significantly associated with response to trastuzumab, TTP, OS and CNS metastases incidence. However, HER2 status determined by IHC and/or FISH assays may not be sufficient to predict response to trastuzumab-based therapy.

Primary signet-ring cell carcinoma of the urinary bladder is a rare histologic variant of adenocarcinoma. Generally, this neoplasm occurs in middle age and the clinical presentation does not differ from the most frequent transitional cell carcinomas. The prognosis is frequently poor as at diagnosis it is often in an advanced phase. It is essential to distinguish this carcinoma from metastases, as different therapeutic strategies are often necessary. We present 5 cases of primary signet-ring cell carcinoma of the urinary bladder and we used a panel of histochemical and immunohistochemical markers for differential diagnosis from secondary carcinoma in an attempt to elucidate the histogenetic derivation of this neoplasia.