Frequent kinase fusions have been reported in spitzoid neoplasms, approximately 10% of which involve ALK rearrangements. Herein, we report a case of atypical Spitz tumor with a novel MLPH-ALK fusion, which has not been previously reported to contribute to cancer development. The tumor was detected in the right arm of a 40-year-old woman. The novel ALK fusion was identified by a 5'-rapid amplification of cDNA ends-based system optimized for formalin-fixed, paraffin-embedded tissue. Initially, ALK expression was detected by immunohistochemistry using 5A4 antibodies for both sensitive and conventional polymer detection methods. However, the anti-ALK1 antibody, which is commonly used for the diagnosis of ALK-positive anaplastic large cell lymphoma, failed to confirm ALK expression. These results indicated that ALK immunohistochemistry results in ALK-rearranged atypical Spitz tumor may differ based on the type of primary antibody clone, which can be a potential diagnostic pitfall.

The use of anaplastic lymphoma kinase antibodies (ALK1) as a diagnostic aid has expanded since becoming a routinely available immunohistochemical stain. Because the skin may be the site of a wide variety of hematolymphoid and fibroblastic proliferations, dermatopathologists commonly use ALK1 as part of a broader staining panel in diagnosing soft tissue and cutaneous hematolymphoid neoplasms. Furthermore, new entities and differential diagnostic contexts are emerging, which broaden the utility of ALK1 immunohistochemistry. We review the expanding role of ALK1 immunohistochemistry in contemporary dermatopathology.

Intravascular large B-cell lymphoma (IVLBCL) is a very rare type of non-Hodgkin lymphoma, usually affecting elderly patients and characterized by selective infiltration of neoplastic cells within blood vessels' lumina. IVLBCL diagnosed with prostatic involvement is extremely rare. We report a patient of 65 years old, having mostly neurological complaints but diagnosed with IVLBCL upon histopathological examination of transurethral prostate resection material, which revealed large neoplastic cell infiltration totally limited within the lumens of small vessels. By immunohistochemistry, neoplastic cell infiltration was positive with MUM1, bcl-6, and bcl-2 and negative with ALK1, CD10, and CD30, with a high Ki-67 proliferation index. CD34 and CD31 staining showed expression in endothelial cells, highlighting the intravascular nature of neoplastic infiltrate. The patient unfortunately refused to receive treatment and died of the disease 8 months after the diagnosis. IVLBCL, though very rare, should be considered in differential diagnosis of all organ biopsies with intravascular infiltration. Further improvements in the understanding of the pathogenesis and biology of this rare type of lymphoma are mandatory.

We describe the case of a 59-year-old woman with Ki-1-negative, T cell-type, anaplastic lymphoma kinase (ALK)-positive large cell lymphoma that was positive for epithelial membrane antigen. She was histopathologically diagnosed as having a metastatic undifferentiated carcinoma from a cervical lymph node biopsy. Clinical and radiographic studies revealed no other primary tumor. The patient underwent a left radical neck dissection. Surgically resected lymph nodes revealed an ALK1-positive large cell lymphoma. Thereafter the patient has had an unusually favorable prognosis.

ALCL is widely recognized with its broad morphologic and phenotypic spectrum causing controversy in the diagnosis of this peculiar neoplasm. It is now beyond doubt that a significant proportion(64 to 84%) of the cases diagnosed as ALCL is closely associated with the expression of chimeric NPM-ALK protein activated by the (2;5) (p23;q35) chromosomal translocation, which can be detected by anti-p80NPM/ALK or ALK1 antibodies. Recently, some investigators including us asserted that these p80NPM/ALK or ALK1-positive(p80/ALK+) ALCLs represent a distinct genetic entity with occurrence in young patients and a favorable prognosis, and should be differentiated from the p80/ALK- tumors with the relatively aggressive clinical course. The p80/ALK+ lymphomas also revealed the characteristic morphology such as horseshoe-like, kidney-like or doughnut-like nuclei and frequent expression of EMA and cytotoxic molecules. However, these features are shared, though to a lesser degree, by other p80/ALK-negative lymphoid neoplasms. Indeed, it is indicated that cytotoxic ALCL cases may be either p80/ALK positive or negative, suggesting that the cytotoxicity and expression of p80/ALK are independent phenomena among the cases of ALCL of T- and null-cell type. Thus, several areas of disagreement and controversy that surround the diagnosis and categorization of ALCL remain.

OBJECTIVE

To explore the clinicopathologic features of CD30-positive sinusoidal large B-cell lymphoma (CD30 + SLBCL) and its relative correlation with Epstein-Barr virus (EBV).

METHODS

Two cases of CD30 + SLBCL, a 65-year-old men and a 85-year-old women were morphologically and immunophenotypically analyzed. EBV status was also evaluated through not only the polymerase chain reaction (PCR) amplification to the EBV Bam HIW DNA sequence, but also an immunohistochemical detection of the latent membrane protein 1 (LMP1).

RESULTS

The patients presented with similarly superficial lymphadenopathy. One of them died of the tumor within 10 months. Microscopically, both of the neoplasms were characterized by a cohesive sinus growth pattern and the monomorphic cytology of the tumor cells. Immunohistochemically, They were both positive for CD45, CD30, and CD20 or CD79alpha, whereas neither expressed EMA, ALK1, nor any histiocytic/T-lineage markers. No evidence of EBV-infection could be found either.

CONCLUSIONS

CD30 + SLBCL is a morphologically and immunophenotypically distinctive variant of diffuse large B-cell lymphoma, which should be distinguished from T/null cell type anaplastic large cell lymphoma and some other nodal lesions with a predominantly sinusoidal infiltrative pattern. CD30 + SLBCL may not be correlation with EBV.

T-cell lineage lymphoma with an intense membranous and paranuclear CD30 expression in the absence of ALK1 raises a differential diagnosis of peripheral T-cell lymphoma (PTCL), NOS and anaplastic large cell lymphoma (ALCL), ALK negative. However, Epstein-Barr virus is consistently negative in ALCL and is not considered an implicating factor in its pathogenesis. We describe a case of T-cell lymphoma showing anaplastic large cell morphology with scattered hallmark cells and a uniform CD30 and Epstein-Barr virus encoded early RNA (EBER) expression that primarily involved the subcutaneous tissue at presentation. On incisional biopsy, the neoplastic cells were positive for CD3, CD2, and CD30 while negative for LCA, CD20, PAX5, CD56, ALK1, and cytotoxic granules. Molecular analysis identified a positive T-cell receptor (beta and gamma) gene rearrangement by PCR. Proliferation index approached 100% and the patient had a rapidly progressive course; the subcutaneous lesions more than doubled in size within couple of weeks with new evidence for widespread systemic involvement. This case emphasizes a rare EBV association with a CD30 positive T-cell lymphoma where the morphologic and immunophenotypic findings are otherwise nondiscriminatory between PTCL, NOS and ALCL, ALK negative.

Inflammatory myofibroblastic tumors (IMTs) are rare mesenchymal tumors initially described in the lung. About half of them exhibit expression of the ALK1 protein, generally resulting from a gene rearrangement. Paranasal sinus IMTs are extremely uncommon, and gene rearrangement of ALK1 is very rare in this localization. A 47-year-old woman presented with rapidly progressive vision loss in her left eye. Clinical and imaging work-up revealed a tumor invading the left ethmoidal and sphenoidal sinuses and extending into the nasal cavity, the orbit and the skull base. Complete tumor resection was performed using an endonasal approach. Pathological examination revealed a paranasal localization of IMT, positive for ALK1 immunostaining. FISH analysis showed an ALK1 gene rearrangement. This case illustrates the local aggressive potential for IMTs. Treatment is primarily surgical, but targeted therapies (crizotinib) might be a solution for ALK1 rearranged cases with a poor prognosis.

OBJECTIVE

To analyze the prevalence of lymphoma subtypes in Shanxi according to the latest World Health Organization (WHO) classification, and to compare the figures with those in other parts of the world.

METHODS

The hematoxylin and eosin-stained sections of 447 lymphoma cases from the archive files of Shanxi Tumor Hospital were reviewed. Immunohistochemical study was performed using a panel of antibodies, including ALK1, bcl-6, CD (1a, 3, 4, 5, 7, 8, 10, 15, 20, 23, 30, 43, 56, 68, 79a and 99), cyclin D1, EMA, IgD, kappa, lambda, LMP1, PAX5, TdT and Vs38C. In addition, in-situ hybridization for Epstein-Barr virus-encoded RNA (EBER) was carried out. All cases were then reclassified according to the latest WHO classification of lymphoma.

RESULTS

Of the 447 cases studied, 385 cases (86.1%) were confirmed to be non-Hodgkin lymphoma (NHL), while 62 cases (13.9%) belonged to classic Hodgkin lymphoma (HL). Of the NHL cases, 68.3% were of B-cell lineage and 30.6% were of T and/or NK-cell lineage. Histiocytic neoplasm accounted for only 0.8% (3 cases). As for the subtyping of NHL, diffuse large B-cell lymphoma was commonest (35.1%), followed by peripheral T-cell lymphoma, NOS (12.0%), extranodal marginal zone B-cell lymphoma (MALT lymphoma) (11.7%), follicular lymphoma (8.6%), T-lymphoblastic lymphoma (7.0%), anaplastic large cell lymphoma (4.2%), B-small lymphocytic lymphoma (3.6%) and mantle cell lymphoma (2.6%). Amongst the 263 cases of B-cell lymphoma, 105 cases (39.9%) expressed immunoglobulin light chain (kappa in 52 cases and lambda in 53 cases) in paraffin sections. Regarding markers for EB virus infection, 14 cases of the B-cell lymphoma gave positive findings with both EBER in-situ hybridization and LMP-1 immunohistochemistry, while 6 of the T/NK-cell lymphoma expressed LMP-1 and 19 showed positive signals for EBER. In NHL, there was discordance in EBER in-situ hybridization and LMP-1 immunohistochemical results. As for HL, EB virus positivity was noted in 37 of the 62 cases (59.7%), including 7 cases of lymphocyte-rich HL, 11 cases of mixed cellularity HL and 19 cases of nodular sclerosis HL. In classic HL, there was complete concordance of results by both EBER in-situ hybridization and LMP-1 immunohistochemistry.

CONCLUSIONS

The prevalence of diffuse large B-cell lymphoma in Shanxi is similar to that in America, Australia, Japan and Korea. The incidence of follicular lymphoma however is much lower than America and Australia.

Neurofibromas of the stomach can occur in the course of Recklinghausen's disease. Sporadic gastric neurofibroma appears rarely. This tumour may look like an ulcer and can be a cause of abdominal pain, nausea, and bleeding from the gastrointestinal tract. We reported a 61-year-old women complaining of stomachache for several months. Gastroscopy revealed a tumour with ulceration in the prepyloric part of the stomach. Helicobacter pylori infection was also present. Helicobacter pylori eradication and prolonged treatment of proton pump inhibitors did not decrease the ailments or the size of the tumour. It was not possible to determine the nature and origin of the tumour by carrying out examinations such as endoscopic ultrasound and computed tomography of the abdomen. Only after surgery and histopathological examination with immunohistochemistry was this tumour identified as a neurofibroma. In order to differentiate the tumour the following immunohistochemical examinations were carried out: CD34 (slightly +), CD117 (-), S-100 (+), desmin (-), NSE (+), GFAP (-), SMA (-), bc12 (-), CD99 (-), ALK1 (-), and MiB (1-1.5%). In such cases excision of the tumour is the preferred treatment.

In the n-alkane-assimilating yeast Yarrowia lipolytica, the transcription of ALK1, encoding cytochrome P450, that catalyses n-alkane hydroxylation is activated by a complex composed of Yas1p and Yas2p via a promoter element, ARE1, in response to n-alkanes. An Opi1-family transcription factor, Yas3p, represses the transcription by binding to Yas2p in the nucleus when cultured in glucose-containing medium, but it is localized to the ER, presumably through interaction with acidic phospholipids, phosphatidic acid and/or phospho inositides, when cultured in n-alkane-containing medium. Here, to elucidate the mechanisms regulating the localization of Yas3p, point and deletion mutants of Yas3p were constructed and analysed. The substitution of Trp(360) and Cys(361) by Arg abrogated the localization of Yas3p to the ER and decreased ARE1-mediated transcriptional activation by n-alkane. A Yas3p truncation mutant consisting of residues 259-422 did not bind to acidic phospholipids, but it was localized to the ER in the presence of n-alkane, implying the acidic-phospholipid-independent recruitment of this mutant to the ER in response to n-alkane. The W360R and C361R substitutions in this truncation mutant abolished its localization to the ER. The results suggest that these residues are implicated in the acidic phospholipid-independent interaction of Yas3p to the ER.

Transforming growth factor β1 (TGF-β1) can promote the proliferation and differentiation of intervertebral disc cells and participates in its repair process. However, whether TGF-β1 engages in the process of disc degeneration has not yet been fully elucidated. The present study aimed to investigate the function of high-dose TGF-β1 on the metabolism of nucleus pulposus cells (NPCs). TGF-β1 levels in human degenerative intervertebral disc tissues and tumor necrosis factor (TNF)-α-induced degenerative NPCs were analyzed. Furthermore, NPCs were treated with TGF-β1 and inhibitors of TGF-β1 receptors [ALK tyrosine kinase receptor (ALK) 1 and ALK5] to determine the effect of the receptors in the mediation of NPC degeneration. The NPC state was determined by the components of secretory collagen I/II, tissue inhibitor of metalloproteinase-3 (TIMP-3) and matrix metalloproteinase (MMP)-13. The mRNA expression of Smad1/2/3/5/8, the downstream gene of TGF-β1 mediated by ALK, was also measured. Results showed that TGF-β1 and ALK1 were positively associated with the degree of degeneration of NP or NPCs in vitro, but negatively associated with ALK5. Furthermore, high-doses of TGF-β1 suppressed collagen II, but enhanced collagen I, TIMP-3, MMP-13, ALK1/5 and Smad1/2/3/5/8 expression. ALK5 inhibition induced the suppression of Smad2/3 and aggravated high-dose TGF-β1-induced NPC degeneration, as shown by the reduction in collagen II and increase in collagen I, TIMP-3 and MMP-13. By contrast, ALK1 inhibition resulted in Smad1/5/8 suppression and alleviated high-dose TGF-β1-induced NPC degeneration. Taken together, it was concluded that high-doses of TGF-β1 contributed to the degeneration of NPCs via the upregulation of ALK1 and Smad1/5/8.

Keywords: ALK tyrosine kinase receptor 1/5; Smad; intervertebral disc degeneration; nucleus pulposus cells; transforming growth factor-β1.

We report a case of ALK1-positive anaplastic large cell lymphoma with expression of placental alkaline phosphatase (PLAP) in many tumor cells. Initially, due to the positivity of tumor cells for CD30 and PLAP, lymph node metastasis of a germ cell neoplasm was discussed. Anaplastic large cell lymphomas of T‑cell lineage form a group of rare non-Hodgkin lymphomas with heterogeneous morphological and immunohistochemical appearance. They may imitate other neoplasms, such as large cell B‑cell lymphomas, metastasis of a carcinoma, melanoma, embryonal carcinoma or seminoma, rhabdomyosarcoma and inflammatory myofibroblastic tumor. Only an extended immunohistochemistry panel leads to an accurate diagnosis.

A new virus species, belonging to the family Potyviridae and capable of infecting most of the soybean cultivars grown in Brazil, was collected in Lavras, Minas Gerais, Brazil, and named Soybean yellow shoot virus (SoyYSV). In this study, the complete 9,052-nucleotide genome of SoyYSV was determined and the structural, biological, and molecular properties of the virus were investigated. The SoyYSV genome encoded a single polyprotein that could be subsequently cleaved, generating 11 proteins. The SoyYSV genome shared 49% nucleotide and 36% amino acid sequence identity with Blackberry virus Y. However, the P1 protein of SoyYSV was much smaller and lacked the ALK1 domain characteristic of the genus Brambyvirus. Electron microscopy revealed flexuous filamentous virus particles, 760 to 780 nm in length, and cytoplasmic inclusions typical of those found in plant cells infected with Potyviridae species. In addition to soybean, SoyYSV infected species in the Amaranthaceae, Caricaceae, Fabaceae, and Solanaceae families. Among the most common potyviruses present in Brazil, only SoyYSV induced local necrotic lesions in Carica papaya L. SoyYSV was transmissible by Myzus persicae and Aphis gossypii but lacked the HC-Pro domain required for aphid transmission in other potyviruses. No seed transmission in soybean was observed.

Specific transgene induction in angiogenic blood vessels has been in demand in gene therapies for several cardiovascular and malignant proliferative diseases in which the vasculature is an essential part of the disease process. In addition to improvements of delivery vehicles, promoters specific to angiogenic blood vessels have been sought for driving expression of the therapeutic gene. Earlier, we isolated a mouse activin receptor-like kinase 1 (Alk1; Acvrl1) transcriptional regulatory fragment that specifically induces transgene expression in newly forming and remodeling arteries in tumor and wound-healing lesions. For the purpose of gene therapy, however, this 9.2 kb regulatory fragment is too large to be incorporated into most viral transfer vectors. To bypass this limitation, highly conserved regions within the Alk1 gene were used to generate a shortened regulatory fragment. In transgenic mice, the shortened 4.8 kb Alk1PIB fragment showed comparable specificities in angiogenic and remodeling feeding arteries. In addition, the Alk1PIB fragment in a recombinant adenovirus vector showed transcriptional activity in cultured human iliac arterial endothelial cells (HIAECs). The Alk1PIB fragment may be used to target therapeutic protein expression in the feeding arteries to assist in the regeneration of ischemic tissues or to induce damages to the lesions, such as malignant tumors.

In this study, we investigated the role of OSH6, which encodes a homolog of the oxysterol-binding protein, in the assimilation of n-alkanes in the yeast Yarrowia lipolytica. The deletion mutant of OSH6 showed growth defects on n-alkanes of 10-16 carbons. In the deletion mutant, production of the functional cytochrome P450 was not observed. However, transcription of ALK1, encoding a major P450 belonging to the CYP52 family that plays a critical role in n-alkane hydroxylation, and further translation of its transcript were noted in the deletion mutant as well as in the wild-type strain. The phospholipid composition was altered and, the ratio of phosphatidylserine (PS) was reduced by the deletion of OSH6. Residues involved in the transport of PS and phosphatidylinositol-4-phosphate in Osh6 of Saccharomyces cerevisiae are conserved in Y. lipolytica Osh6p and substitutions of these residues resulted in a defect in the n-alkane assimilation by Y. lipolytica. From these results, we propose a hypothesis that Osh6p provides an ideal endoplasmic reticulum membrane environment for Alk proteins to have a functional conformation via lipid transport activity in Y. lipolytica.

Background: Activin receptor-like kinase 1 (ALK1) is an endothelial transmembrane serine threonine kinase receptor for BMP family ligands that plays a critical role in cardiovascular development and pathology. Loss-of-function mutations in the ALK1 gene cause type 2 hereditary hemorrhagic telangiectasia (HHT), a devastating disorder that leads to arteriovenous malformations (AVMs). Here we show that ALK1 controls endothelial cell polarization against the direction of blood flow and flow-induced endothelial migration from veins through capillaries into arterioles. Methods: Using Cre lines that recombine in different subsets of arterial, capillary-venous or endothelial tip cells, we showed that capillary-venous Alk1 deletion was sufficient to induce AVM formation in the postnatal retina. Results: ALK1 deletion impaired capillary-venous endothelial cell polarization against the direction of blood flow in vivo and in vitro. Mechanistically, ALK1 deficient cells exhibited increased integrin signaling interaction with VEGFR2, which enhanced downstream YAP/TAZ nuclear translocation. Pharmacological inhibition of integrin or YAP/TAZ signaling rescued flow migration coupling and prevented vascular malformations in Alk1 deficient mice. Conclusions: Our study reveals ALK1 as an essential driver of flow-induced endothelial cell migration and identifies loss of flow-migration coupling as a driver of AVM formation in HHT disease. Integrin-YAP/TAZ signaling blockers are new potential targets to prevent vascular malformations in HHT patients.

Keywords: BMP; HHT; Hippo; Integrin; VEGF.

BMP9 is a cytokine involved in the maturation phase of the angiogenic process that signals through its serine/threonine receptor ALK1 and its coreceptor endoglin. In this paper, we explain how BMP9 directs the regulation of endothelial cell proliferation blockage while in turn stimulating protein synthesis. To achieve this, BMP9 promotes SGK1 synthesis and activation through mTORC2 in order to stimulate the mTORC1/S6K/S6 axis. Moreover, BMP9 blocks proliferation also through SGK1 by reducing the activity of the MEK/ERK signalling pathway. Inhibition of SGK1 activity is sufficient to prevent BMP9-mediated inhibition of ERK, leading to an increase in endothelial cell proliferation. Overall, our findings reveal that SGK1 is a key player during angiogenesis, mediating the pro-quiescent and maturation effects of BMP9/ALK1.

Keywords: ALK1; BMP9; ERK; HHT; SGK1; angiogenesis; endothelial cells; protein synthesis.

Inflammatory myofibroblastic tumors (IMT) are rare and poorly understood inflammatory neoplasms. Most commonly occurring in the liver and gastrointestinal tract, cases of bladder involvement have been rarely reported. Bladder IMT (IMTB) generally presents with gross hematuria and can be differentiated from other bladder tumors by expression of anaplastic lymphoma kinase (ALK1). We report the occurrence of an IMTB detected following lower urinary tract reconstruction with bladder augmentation.

Keywords: bladder tumor; inflammatory myofibroblastic tumor (IMTB); pediatric; pseudotumor.

Objectives: Hereditary hemorrhagic telangiectasia (HHT) is an autosomal dominant vascular disorder characterized by recurrent epistaxis, telangiectasias, and visceral arteriovenous malformations (AVMs). Activin A receptor-like type 1 (ACVRL1/ALK1) and Endoglin (ENG) are the principal genes whose mutations cause HHT. A multicenter study to investigate the correlation between genetic variations and clinical outcomes in Korean HHT patients has been lacking.

Methods: Seventy-two members from 40 families suspected of HHT based on symptoms were genetically screened for pathogenic variants in ACVRL1 and ENG. Patients with genetically diagnosed HHT were also evaluated.

Results: In the HHT genetic screening, 42 patients from 24 of the 40 families had genetic variants that met the pathogenic criteria (pathogenic very strong, pathogenic strong, pathogenic moderate, or pathogenic supporting) based on ACMG Standards and Guidelines in either ENG or ACVRL1; 26 from 12 families (50%) in ENG, and 16 from 12 families (50%) in ACVRL1. The diagnostic screening of 42 genetically positive HHT patients based on the Curaçao criteria revealed that 24 patients (57%) were in the definite group, 17 patients (41%) were in the probable group, and 1 patient (2%) was in the unlikely group. Epistaxis was the most common clinical presentation (38/42, 90%), followed by visceral AVMs (24/42, 57%), and telangiectasia (21/42, 50%). Five patients (12%) did not have a family history of HHT clinical symptoms.

Conclusion: Among patients having ACVRL1 or ENG genetic variants, only about half of them could be clinically diagnosed as definite HHT, suggesting that genetic screening is important to confirm the diagnosis.

Keywords: ACVRL1; ENG; Genetic screening; Hereditary hemorrhagic telangiectasia.

OBJECTIVE

To explore the expression of transforming growth factor beta1 (TGFbeta1) and its type I receptors activin-like kinase 1 (ALK1) and ALK5 mRNA in the development of brain arteriovenous malformation (BAVM).

METHODS

The mRNA expressions of TGFbeta1, ALK1and ALK5 were detected with semiquantitative RT-PCR in patients with BAVM.

RESULTS

The expressions of TGFbeta1 and ALK5 mRNA increased significantly in BAVM, and their relative expression quantity were 0.777-/+0.047 and 0.585-/+0.074, respectively. However, ALK1 mRNA expression declined significantlies with a relative expression of 0.173-/+0.044 in comparison with the control group (0.720-/+0.098, P<0.01).

CONCLUSIONS

The balance of TGFbeta1 and its type I receptors ALK1 and ALK5 mRNA expressions may play important role in the development of BAVM.

Transforming growth factor-β1 (TGF-β1) regulates wound healing/regeneration and aging processes. Dental pulp stem cells from human exfoliated deciduous teeth (SHED) are cell sources for treatment of age-related disorders. We studied the effect of TGF-β1 on SHED and related signaling. SHED were treated with TGF-β1 with/without pretreatment/co-incubation by SB431542, U0126, 5Z-7-oxozeaenol or SB203580. Sircol collagen assay, 3-(4,5-Dimethylthiazol-2-yl)-2,5- diphenyl tetrazolium bromide (MTT) assay, alkaline phosphatase (ALP) assay, RT-PCR, western blotting and PathScan phospho-ELISA were used to measure the effects. We found that SHED expressed ALK1, ALK3, ALK5, TGF-RII, betaglycan and endoglin mRNA. TGF-β1 stimulated p-Smad2, p-TAK1, p-ERK, p-p38 and cyclooxygenase-2 (COX-2) protein expression. It enhanced proliferation and collagen content of SHED that were attenuated by SB431542, 5Z-7-oxozeaenol and SB203580, but not U0126. TGF-β1 (0.5-1 ng/ml) stimulated ALP of SHED, whereas 5-10 ng/ml TGF-β1 suppressed ALP. SB431542 reversed the effects of TGF-β1. However, 5Z-7-oxozeaenol, SB203580 and U0126 only reversed the stimulatory effect of TGF-β1 on ALP. Four inhibitors attenuated TGF-β1-induced COX-2 expression. TGF-β1-stimulated TIMP-1 and N-cadherin was inhibited by SB431542 and 5Z-7-oxozeaenol. These results indicate that TGF-β1 affects SHED by differential regulation of ALK5/Smad2/3, TAK1, p38 and MEK/ERK. TGF-β1 and SHED could potentially be used for tissue engineering/regeneration and treatment of age-related diseases.

Keywords: aging-related diseases; alkaline phosphatase; collagen; cyclooxygenase-2; differentiation.

Endothelial-to-mesenchymal transition (EndMT) indispensable in embryogenesis also occurs in several human pathologies. Although transforming growth factor-β (TGFβ) has been demonstrated to induce EndMT, the type-I receptors (ALK-1 and ALK-5) responsible for TGFβ-induced EndMT is unclear. In the current study, we investigated the role of the Akt1 pathway in ALK1 and ALK5 expression regulation in response to TGFβ1 and TGFβ2 in human microvascular endothelial cells (HMECs). Whereas treatment with TGFβ1 and TGFβ2 or Akt1 gene silencing promoted EndMT accompanied by increased ALK5 expression and reduced ALK1 expression accompanied by increased expression of N-cadherin and reduced expression of eNOS in HMECs, treatment with ALK-5 inhibitor (SB431542) blunted these effects. Importantly, the inhibitor of β-catenin (ICG-001) suppressed TGFβ1- and TGFβ2-induced ALK5 expression in both normal and Akt1 deficient HMECs indicating the integral role of Akt1-β-catenin pathway in the regulation of ALK5 expression promoting EndMT.

Keywords: ALK1; ALK5; Akt1; EndMT; TGFβ; β-catenin.

We recently encountered a case of primary pulmonary angiomatoid fibrous histiocytoma (AFH), which was initially misdiagnosed as inflammatory myofibroblastic tumor (IMT) based in part on anaplastic lymphoma kinase (ALK) expression by immunohistochemistry (IHC). Prompted by this experience, we evaluated ALK expression in 11 AFH, 15 IMT, and 11 follicular dendritic cell sarcomas using 3 different antibody clones: D5F3, 5A4, and ALK1. ALK IHC positive cases were analyzed with fluorescence in situ hybridization (FISH) using dual color ALK break-apart probe kit. The majority of AFH cases studied were positive for ALK IHC with at least 1 antibody (9/11 D5F3, 6/9 5A4, 1/9 ALK1), most demonstrating moderate to strong cytoplasmic staining. AFH with positive ALK IHC showed no ALK gene rearrangement by FISH (0/8) with ALK copy number ranging from 1.6 to 2.1. Sixty-seven percent of IMT were ALK positive by IHC (10/15 D5F3, 8/15 5A4, 7/15 ALK1), and 9 of the 10 cases were positive for ALK gene rearrangement by FISH. All follicular dendritic cell sarcomas were negative for ALK by IHC (D5F3 and 5A4). Our results indicate that ALK expression in AFH is common, particularly with the highly sensitive D5F3 and 5A4 antibodies and enhanced detection systems, and may be a potential source of diagnostic confusion with IMT. The underlying mechanism of ALK expression in AFH is unclear, although it does not appear to be from ALK rearrangement or amplification.