Khush Mittal

A micro-RNA signature associated with race, tumor size, and target gene activity in human uterine leiomyomas

Human uterine leiomyomas (ULMs) are the most common neoplasms of women. Many genes are dysregulated in ULMs and some of this dysregulation may be due to abnormal expression of micro‐RNAs (miRNAs). In this study, 55 ULMs and matched myometrium were collected from 41 patients for microarray‐based global miRNA expression analysis. Of 206 miRNAs examined, 45 miRNAs were significantly up‐ or down‐regulated in ULMs in comparison to the matched myometrium (P < 0.001). The top five dysregulated miRNAs in ULMs are the let‐7 family, miR‐21, miR‐23b, miR‐29b, and miR‐197. Four polycistronic clusters of miRNAs were either up‐ or down‐regulated, but not in a mixed pattern, indicative of coordinated regulation of these miRNAs. Significance analysis revealed that subsets of miRNAs were strongly associated with tumor sizes and race. By prediction analysis we identified some important tumorigenic genes previously identified in ULMs that may be targeted by the dysregulated miRNAs. HMGA2 was identified as one of target genes of the let‐7 family of miRNAs and has been found to be suppressed by let‐7 in vitro. (This article contains Supplementary material available at http://www.interscience.wiley.com/jpages/1045‐2257/suppmat.)

Antiproliferative effects by Let-7 repression of high-mobility group A2 in uterine leiomyoma.

High-mobility group A2 (HMGA2) is commonly overexpressed in large leiomyomas. HMGA2 is an important regulator of cell growth, differentiation, apoptosis, and transformation. As a predicted target of Let-7 microRNAs (Let-7s), HMGA2 can be repressed by Let-7s in vitro. MicroRNA profiling analysis revealed that Let-7s were significantly dysregulated in uterine leiomyomas: high in small leiomyomas and lower in large leiomyomas. To evaluate whether Let-7 repression of HMGA2 plays a major role in leiomyomas, we analyzed the molecular relationship of HMGA2 and Let-7s, both in vitro and in vivo. We first characterized that exogenous Let-7 microRNAs could directly repress the dominant transcript of HMGA2, HMGA2a. This repression was also identified for two cryptic HMGA2 transcripts in primary leiomyoma cultures. Second, we found that the endogenous Let-7s were biologically active and played a major role in the regulation of HMGA2. Then, we illustrated that Let-7 repression of HMGA2 inhibited cellular proliferation. Finally, we examined the expression levels of Let-7c and HMGA2 in a large cohort of leiomyomas (n = 120), and we found high levels of Let-7 and low levels of HMGA2 in small leiomyomas, and low levels of Let-7 and high levels of HMGA2 in large leiomyomas. Our findings suggest that the Let-7–mediated repression of HMGA2 mechanism can be an important molecular event in leiomyoma growth. (Mol Cancer Res 2008;6(4):663–73)

Contralateral ovary in unilateral ovarian carcinoma: a search for preneoplastic lesions.

Contralateral ovaries from patients with unilateral ovarian carcinoma were examined and compared to ovaries from age-matched control patients without ovarian carcinoma. The number of inclusion cysts were increased in ovaries from patients with ovarian carcinoma compared to the controls (p < 0.01). In addition, inclusions from cases with ovarian carcinoma showed serous differentiation more frequently than the controls (p < 0.01; odds ratio = 10.0; 95% confidence interval = 1.2-78.1). An age-related increase in the number of inclusion cysts was seen in the study group but not in the control group. These findings support a role of surface inclusion cysts in the genesis of ovarian carcinoma.

Molecular and immunohistochemical evidence for the origin of uterine leiomyosarcomas from associated leiomyoma and symplastic leiomyoma-like areas

It is uncertain whether uterine leiomyosarcoma arises de novo or in preexisting leiomyoma. Leiomyoma-like areas can be seen associated with uterine leiomyosarcoma, raising the possibility of precursor lesions for uterine leiomyosarcoma. In this study, we examined cases of uterine leiomyosarcoma associated with leiomyoma-like areas at the histological, immunohistochemical and DNA level to further evaluate if benign-looking leiomyoma-like and uterine leiomyosarcoma areas are related. Cases of uterine leiomyosarcoma observed at the New York University Medical Center from 1994 to 2007 were reviewed for the presence of leiomyoma-like areas. Of the 26 cases of uterine leiomyosarcoma observed during this period, 18 cases had an associated leiomyoma-like area (five cellular leiomyoma, four symplastic leiomyoma, four cellular and symplastic leiomyoma and five usual type leiomyoma). Sixteen of the 18 cases were examined immunohistochemically for Ki-67, for estrogen receptor, progesterone receptor and for p53. Immunohistochemical profiles were as expected for leiomyoma-like (the mean expression of p53, ER, PR and Ki-67 at 0.3, 63, 75 and 0.6%, respectively), symplastic leiomyoma-like areas (the mean expression of p53, ER, PR and Ki-67 at 0.6, 85, 89 and 5.5%, respectively) and uterine leiomyosarcoma areas (the mean expression of p53, ER, PR and Ki-67 at 52, 38, 39 and 61%, respectively). In six cases, the leiomyoma-like and uterine leiomyosarcoma areas from each case were examined using high-density oligonucleotide array-CGH to determine genetic aberrations in the two areas. Nearly all the genetic aberrations found in leiomyoma-like areas were also found in the corresponding uterine leiomyosarcoma areas. In addition, uterine leiomyosarcoma areas had additional genetic aberrations. The immunohistochemical profiles and genetic aberrations of the examined cases suggest that uterine leiomyosarcoma could arise from the preexisting leiomyoma-like areas that often have a symplastic or cellular morphology.

Application of Immunohistochemistry to Gynecologic Pathology

CONTEXT A large variety of tumors and lesions arise in the female genital tract. Although the majority of these can be correctly recognized on routine hematoxylin-eosin-stained slides, occasional cases present a diagnostic challenge. Immunohistochemical stains are extremely useful in resolving many of these problematic cases. As the knowledge in this area is constantly expanding, it is useful to have this updated information in a review form for easy access. OBJECTIVE To present our current knowledge of immunohistochemistry of the lesions of the female genital tract in a readily accessible form. DATA SOURCES The review is based on previously published articles on this topic. CONCLUSIONS Immunohistochemical stains help in reaching a conclusive diagnosis in a variety of problematic lesions seen in gynecologic pathology. As in any other system, immunohistochemical findings need to be interpreted in light of the clinical history and morphologic findings.

MED12 and HMGA2 mutations: two independent genetic events in uterine leiomyoma and leiomyosarcoma

Recent identification of somatic MED12 mutations in most uterine leiomyomas brings a new venue for the study of the tumorigenesis of leiomyomas. We are particularly interested in the correlation of MED12 and HMGA2 gene products in leiomyomas and leiomyosarcomas with and without MED12 mutations. To address these issues, in this study we examined MED12 mutations in a large cohort of usual type leiomyomas (178 cases) and uterine leiomyosarcomas (32 cases). We found that 74.7% (133/178) of leiomyomas had MED12 mutations, which was consistent with several independent studies. In contrast, only 9.7% (3/32) of leiomyosarcomas harbored MED12 mutations. Expression analysis by western blot and immunohistochemistry revealed that those leiomyomas with complex MED12 mutations had significantly lower protein products than the matched myometrium. Interestingly, most leiomyosarcomas without MED12 mutations also had very low levels of MED12 expression in comparison to the matched myometrium. These findings suggest a potential functional role of MED12 in both benign and malignant uterine smooth muscle tumors. When we further examined HMGA2 expression in all leiomyomas and leiomyosarcomas, we found that HMGA2 overexpression was exclusively present in those leiomyomas with no MED12 mutation, accounting for 10.1% (18/178) of total leiomyomas and 40% (18/45) of non-MED12 mutant leiomyomas. Twenty-five percent (8/32) of leiomyosarcomas had HMGA2 overexpression, and no MED12 mutations were found in HMGA2 positive leiomyosarcoma. These findings strongly suggest that MED12 mutations and HMGA2 overexpression are independent genetic events that occur in leiomyomas, and they may act differently in the tumorigenesis of uterine leiomyomas.

BK virus infection in AIDS

The BK virus (BKV) belongs to the family of the polyoma group, which contains three species: JC, which is responsible for progressive multifocal leukoencephalopathy in acquired immunodeficiency syndrome (AIDS); simian virus 40 (SV40), which is a simian virus of little pathologic significance in humans; and BKV, which is usually not pathogenic and is found in the urine of asymptomatic individuals. Recently BKV has been reported to cause symptomatic infection in renal transplant patients. The authors report a rare case of a 14-year-old boy with AIDS who developed a BKV infection of the lung and kidney that progressed to diffuse alveolar damage and death. The infected type II pneumocytes in the lung and the tubular epithelial cells in the kidney showed large, homogenous purple intranuclear inclusions. The absence of necrosis and destruction made it possible to distinguish BKV infection from herpes simplex. The size of the infected cells and the lack of a halo around the nuclear inclusion helped rule out cytomegalovirus as the cause of infection. Electron microscopy detected the presence of 40-nm intranuclear viral particles compatible with BKV, and in situ hybridization established the diagnosis.

The accuracy of frozen section in the diagnosis of ovarian neoplasms

In a retrospective study to determine the accuracy of frozen section diagnoses in ovarian neoplasms, the results of consecutive frozen section diagnoses of 311 ovarian neoplasms from two institutions, New York University Medical Center and State University of New York Medical Center at Brooklyn, from 1980 through 1989 were compared with the final diagnosis results following extensive sampling on permanent sections. The final diagnosis was assumed to be correct for purposes of this study. Ovarian neoplasms were correctly diagnosed on frozen section as either benign or malignant in 292 patients (accuracy of 93.8%). Frozen section diagnoses were incorrect in 11 patients (3.5%). Frozen section diagnosis was deferred in 8 instances (2.6%). The positive predictive value was 100%. The negative predictive value was 95.3%, specificity 100%, and sensitivity 86%. There were no false positives. Of the 11 false negative frozen section diagnoses, 9 (82%) were due to limited sampling for frozen section. We therefore suggest that careful examination with sampling of any suspicious lesions be carried out at the time of surgery for patients with benign frozen section diagnosis, since this may avoid a second staging laparotomy, if the final diagnosis is malignant.

Estrogen and progesterone receptor expression in endometrial polyps.

Endometrial polyps are a frequent cause of abnormal uterine bleeding, but their pathogenesis is poorly understood. This study was undertaken to investigate if endometrial polyps result from localized overexpression of estrogen receptors (ERs) or reduced expression of progesterone receptors (PRs). Fourteen cases of endometrial polyps, in which normal cycling endometrium was also present on the same slide, were immunostained for ERs and PRs. Percentages of positive cells in glands and stroma for each receptor were subjectively assessed to the nearest 5%. The intensity of staining was recorded on a scale from 1+ to 4+. The level and intensity of staining in polyps were compared with the staining in normal endometrium. Fewer stromal cells in polyps expressed ERs and PRs compared with cycling endometrium (% ER = 55.9 +/- 25.8 vs. 74.3 +/- 25.8, p = 0.03; % PR = 56.1 +/- 28.2 vs. 87.5 +/- 10.1, p = 0.002). Stroma in polyps also had significantly reduced intensity of staining for PRs, but not for ERs (intensity PR = 2.7 +/- 1.4 vs. 3.5 +/- 0.7, p = 0.015; intensity ER = 2.1 +/- 0.7 vs. 2.4 +/- 0.8, p = 0.45). There were no significant differences in expression of ERs and PRs in the endometrial glands in endometrial polyps compared with normal endometrium (% ER = 75.4 +/- 32.5 vs. 70.7 +/- 39.2. p = 0.25; % PR = 79.6 +/- 32.8 vs. 80.4 +/- 34.4, p = 0.8; intensity ER = 2.7 +/- 0.9 vs. 2.4 +/- 1, p = 0.15; intensity PR = 2.9 +/- 1.4 vs. 3.4 +/- 0.7, p = 0.15). We conclude that endometrial polyps may result from a decrease in ER and PR expression in stromal cells. Because of these receptor-negative stromal cells, endometrial polyps may be relatively insensitive to cyclic hormonal changes.

Proliferating cell nuclear antigen (cyclin) expression in normal and abnormal cervical squamous epithelia.

Expression of Proliferating cell nuclear antigen (PCNA) was evaluated in formalin-fixed, paraffin-embedded normal and abnormal cervical squamous epithelia using immunoperoxidase stains and PC10 monoclonal antibody to PCNA. PCNA was exclusively expressed in the parabasal and basal layers of normal ectocervix and a similar pattern was seen in nine of the 11 cases with squamous metaplasia. Examples of cervical dysplasia showed expression in higher layers of cervical epithelium, corresponding to the degree of dysplasia. Increased staining was seen in condylomas and markedly reduced staining with atrophy. The percentage of basal cells that stained increased progressively from atrophic to normal, to condylomatous, to dysplastic epithelia. Proliferative activity can be satisfactorily assessed in formalin-fixed cervical epithelia using PC10 PCNA antibody. This assessment can be of potential diagnostic use in difficult cases.