Which adults aged 65 and older are at low-risk for cervical spine injuries after low-level falls?

OBJECTIVES: The population is aging and falls are a common reason for emergency department visits. Appropriate imaging in this population is important. The objectives of this study were to estimate the prevalence of cervical spine injury and identify factors associated with cervical spine injuries in adults ≥ 65 years after low-level falls. METHODS: This was a pre-specified sub-study of a prospective observational cohort study of intracranial bleeding in emergency patients ≥ 65 years presenting after low-level falls. The primary outcome was cervical spine injury. The risk factors of interest were Glasgow coma scale (GCS) < 15, head injury, neck pain, age, and frailty defined as Clinical Frailty Scale ≥ 5. Multivariable logistic regression was used to measure the strength of association between risk factors and cervical spine injury. A descriptive analysis of absence of significant risk factors was performed to determine patients who may not require imaging. RESULTS: There were 4308 adults ≥ 65 who sustained low-level falls with mean age of 82.0 (standard deviation ± 8.8) years and 1538 (35.7%) were male; 23 [0.5% (95% confidence interval (CI) 0.3-0.8%)] were diagnosed with cervical spine injuries. The adjusted odds ratios and 95% CIs were 1.3 (0.5-3.2) for GCS < 15, 5.3 (1.7-26.7) for head injury, 13.0 (5.7-31.2) for new neck pain, 1.4 (1.0-1.8) for 5-year increase in age, and 1.1 (0.4-2.9) for frailty. Head injury or neck pain identified all 23 cervical spine injuries. Management was a rigid collar in 19/23 (82.6%) patients and none had surgery. CONCLUSIONS: In emergency patients ≥ 65 years presenting after a low-level fall, head injury, neck pain, and older age were associated with the diagnosis of cervical spine injury. There were no cervical spine injuries in those without head injury or neck pain. Patients with no head injury or neck pain may not require cervical spine imaging.

RéSUMé: OBJECTIFS: La population vieillit et les chutes sont une raison courante pour les visites à l'urgence. Il est important d'avoir une imagerie appropriée dans cette population. Les objectifs de cette étude étaient d'estimer la prévalence des lésions de la colonne cervicale et d'identifier les facteurs associés aux lésions de la colonne cervicale chez les adultes de plus de 65 ans après des chutes de faible niveau. MéTHODES: Il s'agissait d'une sous-étude pré-spécifiée d'une étude prospective de cohorte observationnelle de saignements intracrâniens chez des patients d'urgence de plus de 65 ans se présentant après des chutes de faible niveau. Le résultat principal était une lésion de la colonne cervicale. Les facteurs de risque d'intérêt étaient l'échelle de coma de Glasgow (GCS)<15, les blessures à la tête, les douleurs au cou, l'âge et la fragilité définis comme l'échelle de fragilité clinique >5. La régression logistique multivariée a été utilisée pour mesurer la force de l'association entre les facteurs de risque et les lésions de la colonne cervicale. Une analyse descriptive de l'absence de facteurs de risque significatifs a été réalisée pour déterminer les patients qui ne nécessitent pas d'imagerie. RéSULTATS: Il y avait 4308 adultes de plus de 65 ans qui ont subi des chutes de faible intensité avec un âge moyen de 82.0 ans (écart-type 8.8) et 1538 ans (35.7 %) étaient des hommes; 23 (0.5 % (intervalle de confiance à 95 % 0.3­0.8 %) ont reçu un diagnostic de lésions du rachis cervical. Les rapports de cotes ajustés et les IC à 95 % étaient de 1.3 (0.5­3.2) pour les SCM<15, 5.3 (1.7­26.7) pour les blessures à la tête, 13.0 (5.7­31.2) pour les nouvelles douleurs au cou, 1.4 (1.0 ­ 1.8) pour l'augmentation de l'âge de cinq ans et 1.1 (0.4­2.9) pour la fragilité. Des blessures à la tête ou des douleurs au cou ont permis de déceler les 23 blessures à la colonne cervicale. La prise en charge était un collier rigide chez 19 patients sur 23 (82.6 %) et aucun n'a subi de chirurgie. CONCLUSIONS: Chez les patients d'urgence de plus de 65 ans se présentant après une chute de faible intensité, des blessures à la tête, des douleurs au cou et un âge plus avancé ont été associés au diagnostic de lésion de la colonne cervicale. Il n'y avait pas de blessures à la colonne cervicale chez les personnes sans blessure à la tête ou douleur au cou. Les patients sans blessure à la tête ou douleur au cou peuvent ne pas avoir besoin d'imagerie de la colonne cervicale.

RhoU forms homo-oligomers to regulate cellular responses.

RhoU is an atypical member of the Rho family of small G-proteins, which has N- and C-terminal extensions compared to the classic Rho GTPases RhoA, Rac1 and Cdc42, and associates with membranes through C-terminal palmitoylation rather than prenylation. RhoU mRNA expression is upregulated in prostate cancer and is considered a marker for disease progression. Here, we show that RhoU overexpression in prostate cancer cells increases cell migration and invasion. To identify RhoU targets that contribute to its function, we found that RhoU homodimerizes in cells. We map the region involved in this interaction to the C-terminal extension and show that C-terminal palmitoylation is required for self-association. Expression of the isolated C-terminal extension reduces RhoU-induced activation of p21-activated kinases (PAKs), which are known downstream targets for RhoU, and induces cell morphological changes consistent with inhibiting RhoU function. Our results show for the first time that the activity of a Rho family member is stimulated by self-association, and this is important for its activity.

HDAC Inhibition Restores Response to HER2-Targeted Therapy in Breast Cancer via PHLDA1 Induction.

The downregulation of Pleckstrin Homology-Like Domain family A member 1 (PHLDA1) expression mediates resistance to targeted therapies in receptor tyrosine kinase-driven cancers. The restoration and maintenance of PHLDA1 levels in cancer cells thus constitutes a potential strategy to circumvent resistance to inhibitors of receptor tyrosine kinases. Through a pharmacological approach, we identify the inhibition of MAPK signalling as a crucial step in PHLDA1 downregulation. Further ChIP-qPCR analysis revealed that MEK1/2 inhibition produces significant epigenetic changes at the PHLDA1 locus, specifically a decrease in the activatory marks H3Kme3 and H3K27ac. In line with this, we show that treatment with the clinically relevant class I histone deacetylase (HDAC) inhibitor 4SC-202 restores PHLDA1 expression in lapatinib-resistant human epidermal growth factor receptor-2 (HER2)+ breast cancer cells. Critically, we show that when given in combination, 4SC-202 and lapatinib exert synergistic effects on 2D cell proliferation and colony formation capacity. We therefore propose that co-treatment with 4SC-202 may prolong the clinical efficacy of lapatinib in HER2+ breast cancer patients.

Assembly of nuclear dimers of PI3K regulatory subunits is regulated by the Cdc42-activated tyrosine kinase ACK.

Activated Cdc42-associated kinase (ACK) is an oncogenic nonreceptor tyrosine kinase associated with poor prognosis in several human cancers. ACK promotes proliferation, in part by contributing to the activation of Akt, the major effector of class 1A phosphoinositide 3-kinases (PI3Ks), which transduce signals via membrane phosphoinositol lipids. We now show that ACK also interacts with other key components of class 1A PI3K signaling, the PI3K regulatory subunits. We demonstrate ACK binds to all five PI3K regulatory subunit isoforms and directly phosphorylates p85α, p85ß, p50α, and p55α on Tyr607 (or analogous residues). We found that phosphorylation of p85ß promotes cell proliferation in HEK293T cells. We demonstrate that ACK interacts with p85α exclusively in nuclear-enriched cell fractions, where p85α phosphorylated at Tyr607 (pTyr607) also resides, and identify an interaction between pTyr607 and the N-terminal SH2 domain that supports dimerization of the regulatory subunits. We infer from this that ACK targets p110-independent p85 and further postulate that these regulatory subunit dimers undertake novel nuclear functions underpinning ACK activity. We conclude that these dimers represent a previously undescribed mode of regulation for the class1A PI3K regulatory subunits and potentially reveal additional avenues for therapeutic intervention.

Short-term tetrabromobisphenol A exposure promotes fibrosis of human uterine fibroid cells in a 3D culture system through TGF-beta signaling.

Tetrabromobisphenol A (TBBPA), a derivative of BPA, is a ubiquitous environmental contaminant with weak estrogenic properties. In women, uterine fibroids are highly prevalent estrogen-responsive tumors often with excessive accumulation of extracellular matrix (ECM) and may be the target of environmental estrogens. We have found that BPA has profibrotic effects in vitro, in addition to previous reports of the in vivo fibrotic effects of BPA in mouse uterus. However, the role of TBBPA in fibrosis is unclear. To investigate the effects of TBBPA on uterine fibrosis, we developed a 3D human uterine leiomyoma (ht-UtLM) spheroid culture model. Cell proliferation was evaluated in 3D ht-UtLM spheroids following TBBPA (10-6 -200 µM) administration at 48 h. Fibrosis was assessed using a Masson's Trichrome stain and light microscopy at 7 days of TBBPA (10-3  µM) treatment. Differential expression of ECM and fibrosis genes were determined using RT² Profiler™ PCR arrays. Network and pathway analyses were conducted using Ingenuity Pathway Analysis. The activation of pathway proteins was analyzed by a transforming growth factor-beta (TGFB) protein array. We found that TBBPA increased cell proliferation and promoted fibrosis in 3D ht-UtLM spheroids with increased deposition of collagens. TBBPA upregulated the expression of profibrotic genes and corresponding proteins associated with the TGFB pathway. TBBPA activated TGFB signaling through phosphorylation of TGFBR1 and downstream effectors-small mothers against decapentaplegic -2 and -3 proteins (SMAD2 and SMAD3). The 3D ht-UtLM spheroid model is an effective system for studying environmental agents on human uterine fibrosis. TBBPA can promote fibrosis in uterine fibroid through TGFB/SMAD signaling.

Statins change the cytokine profile in Trypanosoma cruzi-infected U937 macrophages and murine cardiac tissue through Rho-associated kinases inhibition.

Introduction: Chronic Chagasic cardiomyopathy (CCC), caused by the protozoan Trypanosoma cruzi, is the most severe manifestation of Chagas disease.CCC is characterized by cardiac inflammation and fibrosis caused by a persistent inflammatory response. Following infection, macrophages secrete inflammatory mediators such as IL-1ß, IL-6, and TNF-α to control parasitemia. Although this response contains parasite infection, it causes damage to the heart tissue. Thus, the use of immunomodulators is a rational alternative to CCC. Rho-associated kinase (ROCK) 1 and 2 are RhoA-activated serine/threonine kinases that regulate the actomyosin cytoskeleton. Both ROCKs have been implicated in the polarization of macrophages towards an M1 (pro-inflammatory) phenotype. Statins are FDA-approved lipid-lowering drugs that reduce RhoA signaling by inhibiting geranylgeranyl pyrophosphate (GGPP) synthesis. This work aims to identify the effect of statins on U937 macrophage polarization and cardiac tissue inflammation and its relationship with ROCK activity during T. cruzi infection. Methods: PMA-induced, wild-type, GFP-, CA-ROCK1- and CA-ROCK2-expressing U937 macrophages were incubated with atorvastatin, or the inhibitors Y-27632, JSH-23, TAK-242, or C3 exoenzyme incubated with or without T. cruzi trypomastigotes for 30 min to evaluate the activity of ROCK and the M1 and M2 cytokine expression and secretion profiling. Also, ROCK activity was determined in T. cruzi-infected, BALB/c mice hearts. Results: In this study, we demonstrate for the first time in macrophages that incubation with T. cruzi leads to ROCK activation via the TLR4 pathway, which triggers NF-κB activation. Inhibition of ROCKs by Y-27632 prevents NF-κB activation and the expression and secretion of M1 markers, as does treatment with atorvastatin. Furthermore, we show that the effect of atorvastatin on the NF-kB pathway and cytokine secretion is mediated by ROCK. Finally, statin treatment decreased ROCK activation and expression, and the pro-inflammatory cytokine production, promoting anti-inflammatory cytokine expression in chronic chagasic mice hearts. Conclusion: These results suggest that the statin modulation of the inflammatory response due to ROCK inhibition is a potential pharmacological strategy to prevent cardiac inflammation in CCC.

Targeting Rho GTPase Signaling Networks in Cancer.

As key regulators of cytoskeletal dynamics, Rho GTPases coordinate a wide range of cellular processes, including cell polarity, cell migration, and cell cycle progression. The adoption of a pro-migratory phenotype enables cancer cells to invade the stroma surrounding the primary tumor and move toward and enter blood or lymphatic vessels. Targeting these early events could reduce the progression to metastatic disease, the leading cause of cancer-related deaths. Rho GTPases play a key role in the formation of dynamic actin-rich membrane protrusions and the turnover of cell-cell and cell-extracellular matrix adhesions required for efficient cancer cell invasion. Here, we discuss the roles of Rho GTPases in cancer, their validation as therapeutic targets and the challenges of developing clinically viable Rho GTPase inhibitors. We review other therapeutic targets in the wider Rho GTPase signaling network and focus on the four best characterized effector families: p21-activated kinases (PAKs), Rho-associated protein kinases (ROCKs), atypical protein kinase Cs (aPKCs), and myotonic dystrophy kinase-related Cdc42-binding kinases (MRCKs).

Previously Diagnosed Reticulum Cell Hyperplasia in Decalcified Rat Bone Marrow Stain Positive for Ionized Calcium Binding Adapter Molecule 1 (Iba1): A Monocytic/Macrophage Cell Marker.

Reticulum cell hyperplasia (RCH) was a term used for many years by the National Toxicology Program (NTP) to describe a certain non-neoplastic bone marrow lesion of rats. Retrospective microscopic evaluation of RCH lesions and immunohistochemistry analyses were performed to reassess and further characterize these lesions. The NTP database was searched to identify femoral bone marrow specimens diagnosed with RCH from 1981 to 2014 (n = 254). The diagnosis last occurred in 2003, after which the term "cellular infiltration" was used. Eighty-three RCH slides, spanning 22 years, representing 34 different chemicals, were selected for microscopic review, and a subset (23) was chosen for ionized calcium binding adapter molecule 1 (Iba1) immunohistochemical staining; initial investigations revealed Iba1 worked as a macrophage marker on decalcified tissue. The following diagnoses were made upon reevaluation: 36 were consistent with cellularity increased, macrophage, 22 with histiocytic sarcoma, 8 with increased myeloid cells, 4 with autolysis, and 13 were normal appearance. All 23 RCH lesions stained positive for Iba1. Fifty-eight of 83 bone marrows previously diagnosed with RCH are consistent morphologically and immunohistochemically with cells of histiocytic origin. These results will help with interpretation of historical data and demonstrates that Iba1 can be used in decalcified bone marrow sections.

Emerging Roles of Fibroblast Growth Factor 10 in Cancer.

Whilst cross-talk between stroma and epithelium is critical for tissue development and homeostasis, aberrant paracrine stimulation can result in neoplastic transformation. Chronic stimulation of epithelial cells with paracrine Fibroblast Growth Factor 10 (FGF10) has been implicated in multiple cancers, including breast, prostate and pancreatic ductal adenocarcinoma. Here, we examine the mechanisms underlying FGF10-induced tumourigenesis and explore novel approaches to target FGF10 signaling in cancer.

The Life Cycle of the Uterine Fibroid Myocyte.

PURPOSE OF REVIEW: Uterine fibroids are common benign tumors of women in the USA and worldwide, yet the biological nature and pathogenesis of these tumors remain largely unknown. This review presents our view of the stages in the life cycle of a subset of uterine fibroid myocytes, introduces hypothetical concepts and morphological data to explain these changes, and relates these changes in individual myocytes to the phases of fibroid tumor development. RECENT FINDINGS: The observations gained from light and electron microscopic, immunohistochemical, and morphometric studies in our laboratory have led to the hypothesis that fibroid changes over time may relate to the excessive production of collagen by phenotypically transformed myocytes. This accumulation of collagen results in decreased microvessel density, followed by myocyte injury and atrophy, with eventual senescence and involution through ischemic cellular degeneration and inanition. SUMMARY: Uterine leiomyomas, or fibroids, are characterized by two histologic features-proliferation of myocytes and production of an extracellular collagenous matrix. In the larger tumors, the collagenous matrix is often abundant. Within those regions in which the accumulating collagen is excessive, the myocytes are progressively separated from their blood supply, resulting in myocyte atrophy and eventually cell death. It is within these hypocellular, hyalinized areas that the complete lifecycle of the fibroid myocyte is realized. It begins with the phenotypic transformation of a contractile cell to one characterized by proliferation and collagen synthesis, progresses through an intermediate stage of atrophy related to interstitial ischemia, and eventuates in cell death due to inanition. Lastly, resorption of inanotic cells appears to occur by a non-phagocytic, presumably enzymatic process of degradation and recycling that we refer to as reclamation.

Preparation of Three-dimensional (3-D) Human Liver (HepaRG) Cultures for Histochemical and Immunohistochemical Staining and Light Microscopic Evaluation.

The use of three-dimensional (3-D) in vitro culture systems (spheroids, organoids) in biomolecular and drug discovery research has become increasingly popular. The popularity is due, in part, to a diminished reliance on animal bioassays and a desire to develop physiologically relevant cell culture systems that simulate the in vivo tissue microenvironment. Most evaluations of 3-D cultures are by confocal microscopy and high-content imaging; however, these technologies do not allow for detailed cellular morphologic assessments or permit basic hematoxylin and eosin histologic evaluations. There are few studies that have reported detailed processes for preparing 3-D cultures for paraffin embedding and subsequent use for histochemical or immunohistochemical staining. In an attempt to do so, we have developed a protocol to paraffin-embed human liver spheroids that can be sectioned with a microtome and mounted onto glass slides for routine histochemical and immunohistochemical staining and light microscopic evaluations.

PHLDA1 Mediates Drug Resistance in Receptor Tyrosine Kinase-Driven Cancer.

Development of resistance causes failure of drugs targeting receptor tyrosine kinase (RTK) networks and represents a critical challenge for precision medicine. Here, we show that PHLDA1 downregulation is critical to acquisition and maintenance of drug resistance in RTK-driven cancer. Using fibroblast growth factor receptor (FGFR) inhibition in endometrial cancer cells, we identify an Akt-driven compensatory mechanism underpinned by downregulation of PHLDA1. We demonstrate broad clinical relevance of our findings, showing that PHLDA1 downregulation also occurs in response to RTK-targeted therapy in breast and renal cancer patients, as well as following trastuzumab treatment in HER2+ breast cancer cells. Crucially, knockdown of PHLDA1 alone was sufficient to confer de novo resistance to RTK inhibitors and induction of PHLDA1 expression re-sensitized drug-resistant cancer cells to targeted therapies, identifying PHLDA1 as a biomarker for drug response and highlighting the potential of PHLDA1 reactivation as a means of circumventing drug resistance.

Inhibition of Ral GTPases Using a Stapled Peptide Approach.

Aberrant Ras signaling drives numerous cancers, and drugs to inhibit this are urgently required. This compelling clinical need combined with recent innovations in drug discovery including the advent of biologic therapeutic agents, has propelled Ras back to the forefront of targeting efforts. Activated Ras has proved extremely difficult to target directly, and the focus has moved to the main downstream Ras-signaling pathways. In particular, the Ras-Raf and Ras-PI3K pathways have provided conspicuous enzyme therapeutic targets that were more accessible to conventional drug-discovery strategies. The Ras-RalGEF-Ral pathway is a more difficult challenge for traditional medicinal development, and there have, therefore, been few inhibitors reported that disrupt this axis. We have used our structure of a Ral-effector complex as a basis for the design and characterization of α-helical-stapled peptides that bind selectively to active, GTP-bound Ral proteins and that compete with downstream effector proteins. The peptides have been thoroughly characterized biophysically. Crucially, the lead peptide enters cells and is biologically active, inhibiting isoform-specific RalB-driven cellular processes. This, therefore, provides a starting point for therapeutic inhibition of the Ras-RalGEF-Ral pathway.

Renal Cell Carcinomas in Vinylidene Chloride-exposed Male B6C3F1 Mice Are Characterized by Oxidative Stress and TP53 Pathway Dysregulation.

Vinylidene chloride (VDC) has been widely used in the production of plastics and flame retardants. Exposure of B6C3F1 mice to VDC in the 2-year National Toxicology Program carcinogenicity bioassay resulted in a dose-dependent increases in renal cell hyperplasia, renal cell adenoma, and renal cell carcinomas (RCCs). Among those differentially expressed genes from controls and RCC of VDC-exposed mice, there was an overrepresentation of genes from pathways associated with chronic xenobiotic and oxidative stress as well as c-Myc overexpression and dysregulation of TP53 cell cycle checkpoint and DNA damage repair pathways in RCC. Trend analysis comparing RCC, VDC-exposed kidney, and chamber control kidney showed a conservation of pathway dysregulation in terms of overrepresentation of xenobiotic and oxidative stress, and DNA damage and cell cycle checkpoint pathways in both VDC-exposed kidney and RCC, suggesting that these mechanisms play a role in the pathogenesis of RCC in VDC-exposed mice.

Gene expression of mesothelioma in vinylidene chloride-exposed F344/N rats reveal immune dysfunction, tissue damage, and inflammation pathways.

A majority (∼80%) of human malignant mesotheliomas are asbestos-related. However, non-asbestos risk factors (radiation, chemicals, and genetic factors) account for up to 30% of cases. A recent 2-year National Toxicology Program carcinogenicity bioassay showed that male F344/N rats exposed to the industrial toxicant vinylidene chloride (VDC) resulted in a marked increase in malignant mesothelioma. Global gene expression profiles of these tumors were compared to spontaneous mesotheliomas and the F344/N rat mesothelial cell line (Fred-PE) in order to characterize the molecular features and chemical-specific profiles of mesothelioma in VDC-exposed rats. As expected, mesotheliomas from control and VDC-exposed rats shared pathways associated with tumorigenesis, including cellular and tissue development, organismal injury, embryonic development, inflammatory response, cell cycle regulation, and cellular growth and proliferation, while mesotheliomas from VDC-exposed rats alone showed overrepresentation of pathways associated with pro-inflammatory pathways and immune dysfunction such as the nuclear factor kappa-light-chain-enhancer of activated B cells signaling pathway, interleukin (IL)-8 and IL-12 signaling, interleukin responses, Fc receptor signaling, and natural killer and dendritic cells signaling, as well as overrepresentation of DNA damage and repair. These data suggest that a chronic, pro-inflammatory environment associated with VDC exposure may exacerbate disturbances in oncogene, growth factor, and cell cycle regulation, resulting in an increased incidence of mesothelioma.

Histopathological and Immunohistochemical Characterization of Methyl Eugenol-induced Nonneoplastic and Neoplastic Neuroendocrine Cell Lesions in Glandular Stomach of Rats.

Methyl eugenol induces neuroendocrine (NE) cell hyperplasia and tumors in F344/N rat stomach. Detailed histopathological and immunohistochemical (IHC) characterization of these tumors has not been previously reported. The objective of this study was to fill that data gap. Archived slides and paraffin blocks were retrieved from the National Toxicology Program Archives. NE hyperplasias and tumors were stained with chromogranin A, synaptophysin, amylase, gastrin, H(+)/K(+) adenosine triphosphatase (ATPase), pepsinogen, somatostatin, and cytokeratin 18 (CK18) antibodies. Many of the rats had gastric mucosal atrophy, due to loss of chief and parietal cells. The hyperplasias and tumors were confined to fundic stomach, and females were more affected than the males. Hyperplasia of NE cells was not observed in the pyloric region. Approximately one-third of the females with malignant NE tumors had areas of pancreatic acinar differentiation. The rate of metastasis was 21%, with liver being the most common site of metastasis. Immunohistochemically, the hyperplasias and tumors stained consistently with chromogranin A and synaptophysin. Neoplastic cells were also positive for amylase and CK18 and negative for gastrin, somatostatin, H(+)/K(+) ATPase, and pepsinogen. Metastatic neoplasms histologically similar to the primary neoplasm stained positively for chromogranin A and synaptophysin. Based on the histopathological and IHC features, the neoplasms appear to arise from enterochromaffin-like cells.

Spontaneous mesotheliomas in F344/N rats are characterized by dysregulation of cellular growth and immune function pathways.

Aged male Fischer 344/N rats are prone to developing spontaneous peritoneal mesotheliomas that arise predominantly from the tunica vaginalis of the testes. A definitive cause for the predominance of this neoplasm in F344/N rats is unknown. Investigation of the molecular alterations that occur in spontaneous rat mesotheliomas may provide insight into their pathogenesis as well enable a better understanding regarding the mechanisms underlying chemically induced mesothelioma in rodents. Mesothelial cell function represents a complex interplay of pathways related to host defense mechanisms and maintenance of cellular homeostasis. Global gene expression profiles of spontaneous mesotheliomas from vehicle control male F344/N rats from 2-year National Toxicology Program carcinogenicity bioassays were analyzed to determine the molecular features of these tumors and elucidate tumor-specific gene expression profiles. The resulting gene expression pattern showed that spontaneous mesotheliomas are associated with upregulation of various growth factors, oncogenes, cytokines, pattern recognition response receptors, and pathogen-associated molecular patterns receptors, and the production of reactive oxygen and nitrogen species, as well as downregulation of apoptosis pathways. Alterations in these pathways in turn trigger molecular responses that stimulate cell proliferation and promote tumor survival and progression.

The natural history of uterine leiomyomas: morphometric concordance with concepts of interstitial ischemia and inanosis.

Based upon our morphologic observations, we hypothesize and also provide morphometric evidence for the occurrence of progressive developmental changes in many uterine fibroids, which can be arbitrarily divided into 4 phases. These developmental phases are related to the ongoing production of extracellular collagenous matrix, which eventually exceeds the degree of angiogenesis, resulting in the progressive separation of myocytes from their blood supply and a condition of interstitial ischemia. The consequence of this process of slow ischemia with nutritional and oxygen deprivation is a progressive myocyte atrophy (or inanition), culminating in cell death, a process that we refer to as inanosis. The studies presented here provide quantitative and semiquantitative evidence to support the concept of the declining proliferative activity as the collagenous matrix increases and the microvascular density decreases.

Characterization of four new mouse cytochrome P450 enzymes of the CYP2J subfamily.

The cytochrome P450 superfamily encompasses a diverse group of enzymes that catalyze the oxidation of various substrates. The mouse CYP2J subfamily includes members that have wide tissue distribution and are active in the metabolism of arachidonic acid (AA), linoleic acid (LA), and other lipids and xenobiotics. The mouse Cyp2j locus contains seven genes and three pseudogenes located in a contiguous 0.62 megabase cluster on chromosome 4. We describe four new mouse CYP2J isoforms (designated CYP2J8, CYP2J11, CYP2J12, and CYP2J13). The four cDNAs contain open reading frames that encode polypeptides with 62-84% identity with the three previously identified mouse CYP2Js. All four new CYP2J proteins were expressed in Sf21 insect cells. Each recombinant protein metabolized AA and LA to epoxides and hydroxy derivatives. Specific antibodies, mRNA probes, and polymerase chain reaction primer sets were developed for each mouse CYP2J to examine their tissue distribution. CYP2J8 transcripts were found in the kidney, liver, and brain, and protein expression was confirmed in the kidney and brain (neuropil). CYP2J11 transcripts were most abundant in the kidney and heart, with protein detected primarily in the kidney (proximal convoluted tubules), liver, and heart (cardiomyocytes). CYP2J12 transcripts were prominently present in the brain, and CYP2J13 transcripts were detected in multiple tissues, with the highest expression in the kidney. CYP2J12 and CYP2J13 protein expression could not be determined because the antibodies developed were not immunospecific. We conclude that the four new CYP2J isoforms might be involved in the metabolism of AA and LA to bioactive lipids in mouse hepatic and extrahepatic tissues.

Immunohistochemical features of 3,3',4,4'-tetrachloroazobenzene-induced rat gingival lesions.

Gingival lesions of squamous hyperplasia, cystic keratinizing hyperplasia (CKH), and squamous cell carcinoma (SCC) can be induced in rats treated by chronic gavage with 10-100 mg/kg 3,3',4,4'-tetrachloroazobenzene. We evaluated gingival squamous hyperplasia (GSH), CKH, and SCC for the immunohistochemical pattern of expression of carcinogenesis-associated markers. The 3 types of lesions and controls were stained with proliferation markers (proliferating cell nuclear antigen [PCNA] and cyclin-D1), tumor-suppressor markers (ß-catenin and mammary serine protease inhibitor [maspin]) and stroma-related markers (α-smooth muscle actin [SMA] and osteonectin/SPARC). The lesions had common immunohistochemical characteristics that differed in their expression patterns among the various diagnoses. PCNA and cyclin-D1 expression was higher in GSH, CKH, and SCC than in controls. The normal membranous expression of ß-catenin was lower in GSH, and almost absent in CKH and SCC. Maspin expression was similar in GSH and controls, whereas both CKH and SCC showed decreased expression. SMA and/or osteonectin/SPARC were seen in stromal cells in CKH and SCC. Collectively, there appears to be a progression from hyperplastic and cystic lesions toward malignancy based on the morphological changes, supported by the expression of carcinogenesis-associated proteins. The exact sequence of events leading to SCC remains to be defined in a time-dependent manner.