NDRG1/2 expression is inhibited in primary acute myeloid leukemia

Expression of N-myc downregulated gene 1 (NDRG1) is associated with growth arrest and differentiation of tumor cells. In hematopoietic cells, NDRG1 was identified in a screen for differentiation-related genes in human myelomonocytic leukemic U937 cells. In the present study, we found significantly higher NDRG1 mRNA levels in granulocytes of healthy donors than in primary acute myeloid leukemia (AML) cells. Another NDRG family member, NDRG2, was significantly higher expressed in normal macrophages compared to primary AML cells. Moreover, NDRG1 mRNA levels increased in two acute promyelocytic leukemia (APL) patients as well as in NB4 and HT93 APL cells upon all-trans retinoic acid (ATRA) therapy. In line with these observations, silencing of NDRG1 diminished neutrophil differentiation of leukemic cell lines. In conclusion, we found an association of low NDRG1 levels with an immature cell phenotype and provide evidence that NDRG1 is functionally involved in neutrophil maturation.

Complexity of miR-223 regulation by CEBPA in human AML

microRNA-223 (miR-223) can trigger normal granulopoiesis. miR-223 expression is regulated by two distinct CEBPA (CCAAT/enhancer binding protein-alpha) sites. Here, we report that miR-223 is largely suppressed in cells from acute myeloid leukemia (AML) patients. By sequencing, we found that miR-223 suppression in AML is not caused by DNA sequence alterations, nor is it mediated by promoter hypermethylation. The analysis of the individual contribution of both CEBPA sites to miR-223 regulation identified the site upstream of the miR-223 primary transcript as the predominant regulatory element. Our results suggest that miR-223 suppression in AML is caused by impaired miR-223 upstream factors.

Heterozygous deletion of the PU.1 locus in human AML

The transcription factor PU.1 is essential for myeloid development. Targeted disruption of an upstream regulatory element (URE) decreases PU.1 expression by 80% and leads to acute myeloid leukemia (AML) in mice. Here, we sequenced the URE sequences of PU.1 in 120 AML patients. Four polymorphisms (single nucleotide polymorphisms [SNPs]) in the URE were observed, with homozygosity in all SNPs in 37 patients. Among them, we compared samples at diagnosis and remission, and one patient with cytogenetically normal acute myeloid leukemia M2 was identified with heterozygosity in 3 of the SNPs in the URE at remission. Loss of heterozygosity was further found in this patient at 2 polymorphic sites in the 5' promoter region and in 2 intronic sites flanking exon 4, thus suggesting loss of heterozygosity covering at least 40 kb of the PU.1 locus. Consistently, PU.1 expression in this patient was markedly reduced. Our study suggests that heterozygous deletion of the PU.1 locus can be associated with human AML.

Unfolded protein response suppresses CEBPA by induction of calreticulin in acute myeloid leukaemia

The unfolded protein response (UPR) is triggered by the accumulation of misfolded proteins within the endoplasmic reticulum (ER). The role of the UPR during leukemogenesis is unknown so far. Here, we studied the induction of mediators of the UPR in leukaemic cells of AML patients. Increased expression of the spliced variant of the X-box binding protein 1 (XBP1s) was detected in 17.4% (16 of 92) of AML patients. Consistent with activated UPR, this group also had increased expression of ER-resident chaperones such as the 78 kD glucose-regulated protein (GRP78) and of calreticulin. Conditional expression of calreticulin in leukaemic U937 cells was found to increase calreticulin binding to the CEBPA mRNA thereby efficiently blocking translation of the myeloid key transcription factor CEBPA and ultimately affecting myeloid differentiation. Consequently, leukaemic cells from AML patients with activated UPR and thus increased calreticulin levels showed in fact suppressed CEBPA protein expression. We identified two functional ER stress response elements (ERSE) in the calreticulin promoter. The presence of NFY and ATF6, as well as an intact binding site for YY1 within these ERSE motifs were essential for mediating sensitivity to ER stress and activation of calreticulin. Thus, we propose a model of the UPR being activated in a considerable subset of AML patients through induction of calreticulin along the ATF6 pathway, thereby ultimately suppressing CEBPA translation and contributing to the block in myeloid differentiation.

Lineage-specific transcription factor aberrations in AML

Transcription factors play a key role in the commitment of hematopoietic stem cells to differentiate into specific lineages [78]. This is particularly important in that a block in terminal differentiation is the key contributing factor in acute leukemias. This general theme of the role of transcription factors in differentiation may also extend to other tissues, both in terms of normal development and cancer. Consistent with the role of transcription factors in hematopoietic lineage commitment is the frequent finding of aberrations in transcription factors in AML patients. Here, we intend to review recent findings on aberrations in lineage-restricted transcription factors as observed in patients with acute myeloid leukemia (AML).

Examination of FGFRL1 as a candidate gene for diaphragmatic defects at chromosome 4p16.3 shows that Fgfrl1 null mice have reduced expression of Tpm3, sarcomere genes and Lrtm1 in the diaphragm

Fgfrl1 (also known as Fgfr5; OMIM 605830) homozygous null mice have thin, amuscular diaphragms and die at birth because of diaphragm hypoplasia. FGFRL1 is located at 4p16.3, and this chromosome region can be deleted in patients with congenital diaphragmatic hernia (CDH). We examined FGFRL1 as a candidate gene for the diaphragmatic defects associated with 4p16.3 deletions and re-sequenced this gene in 54 patients with CDH. We confirmed six known coding single nucleotide polymorphisms (SNPs): c.209G > A (p.Pro20Pro), c.977G > A (p.Pro276Pro), c.1040T > C (p.Asp297Asp), c.1234C > A (p.Pro362Gln), c.1420G > T (p.Arg424Leu), and c.1540C > T (p.Pro464Leu), but we did not identify any gene mutations. We genotyped additional CDH patients for four of these six SNPs, including the three non-synonymous SNPs, to make a total of 200 chromosomes, and found that the allele frequency for the four SNPs, did not differ significantly between patients and normal controls (p > or = 0.05). We then used Affymetrix Genechip Mouse Gene 1.0 ST arrays and found eight genes with significantly reduced expression levels in the diaphragms of Fgfrl1 homozygous null mice when compared with wildtype mice-Tpm3, Fgfrl1 (p = 0.004), Myl2, Lrtm1, Myh4, Myl3, Myh7 and Hephl1. Lrtm1 is closely related to Slit3, a protein associated with herniation of the central tendon of the diaphragm in mice. The Slit proteins are known to regulate axon branching and cell migration, and inhibition of Slit3 reduces cell motility and decreases the expression of Rac and Cdc42, two genes that are essential for myoblast fusion. Further studies to determine if Lrtm1 has a similar function to Slit3 and if reduced Fgfrl1 expression can cause diaphragm hypoplasia through a mechanism involving decreased myoblast motility and/or myoblast fusion, seem indicated.

Type IV delayed-type hypersensitivity of the respiratory tract due to budesonide use: report of two cases and a literature review

Respiratory type-IV hypersensitivity reactions due to corticosteroids is a rare phenomenon. We describe two such cases. The first is a 37- year-old atopic woman who developed labial angioedema and nasal itching after the use of budesonide nasal spray. A month later, after the first puffs of a formoterol/budesonide spray prescribed for asthma, she noticed symptoms of tongue and oropharyngeal itching and redness with subsequent dysphagia, labial and tongue angioedema, and facial oedema. The second is a 15-year-old non-atopic woman who reported pruritic eruptions around the nostrils after using a budesonide nasal spray. A year later she presented with nasal pruritus with intense congestion and labial and facial oedema after using the same spray. Both patients were evaluated with patch-tests using the commercial T.R.U.E. test, a budesonide solution, and corticosteroid creams. Test evaluation was performed at 48 and 96 hours. In both patients, patch tests were positive to budesonide (++) on the second day. The first patient also had a positive (+) reaction to tixocortol-21-pivalate. All the other patch tests were negative. Clinicians should be aware that hypersensitivity reactions may occur during the use of nasal or inhaled corticosteroids.

Processing-dependent and -independent pathways for recognition of iodinated contrast media by specific human T cells

One to three percent of patients exposed to intravenously injected iodinated contrast media (CM) develop delayed hypersensitivity reactions. Positive patch test reactions, immunohistological findings, and CM-specific proliferation of T cells in vitro suggest a pathogenetic role for T cells. We have previously demonstrated that CM-specific T cell clones (TCCs) show a broad range of cross-reactivity to different CM. However, the mechanism of specific CM recognition by T cell receptors (TCRs) has not been analysed so far.

The FGFRL1 receptor is shed from cell membranes, binds fibroblast growth factors (FGFs), and antagonizes FGF signaling in Xenopus embryos

FGFRL1 (fibroblast growth factor receptor like 1) is the fifth and most recently discovered member of the fibroblast growth factor receptor (FGFR) family. With up to 50% amino acid similarity, its extracellular domain closely resembles that of the four conventional FGFRs. Its intracellular domain, however, lacks the split tyrosine kinase domain needed for FGF-mediated signal transduction. During embryogenesis of the mouse, FGFRL1 is essential for the development of parts of the skeleton, the diaphragm muscle, the heart, and the metanephric kidney. Since its discovery, it has been hypothesized that FGFRL1 might act as a decoy receptor for FGF ligands. Here we present several lines of evidence that support this notion. We demonstrate that the FGFRL1 ectodomain is shed from the cell membrane of differentiating C2C12 myoblasts and from HEK293 cells by an as yet unidentified protease, which cuts the receptor in the membrane-proximal region. As determined by ligand dot blot analysis, cell-based binding assays, and surface plasmon resonance analysis, the soluble FGFRL1 ectodomain as well as the membrane-bound receptor are capable of binding to some FGF ligands with high affinity, including FGF2, FGF3, FGF4, FGF8, FGF10, and FGF22. We furthermore show that ectopic expression of FGFRL1 in Xenopus embryos antagonizes FGFR signaling during early development. Taken together, our data provide strong evidence that FGFRL1 is indeed a decoy receptor for FGFs.

Rapid spontaneous malignant progression of supratentorial tanycytic ependymoma with sarcomatous features - "Ependymosarcoma"

By analogy to gliosarcoma, the term "ependymosarcoma" has recently been coined to thematize the rare phenomenon of a malignant mesenchymal component arising within an ependymoma. We report on an example of this paradigm, involving tanycytic ependymoma as the host tumor in a 40-year-old female who underwent two tumor extirpation procedures at one-year interval. She first presented with severe headaches, and was seen by imaging to harbor a moderately enhancing mass 2.5cm in diameter at the rostral septum pellucidum accompanied by occlusive hydrocephalus. Microscopically, the tumor consisted of solid, wavy fascicles of elongated cells that were occasionally interrupted by vague perivascular pseudorosettes. Mitotic activity was absent, and less than 1% of nuclei immunoreacted for MIB-1. A histological diagnosis of tanycytic ependymoma (WHO grade II) was rendered, and no adjuvant therapy given. At recurrence, the lesion was 3.5cm in diameter, intensely enhancing, and had already seeded into the subarachnoid space. Histology showed a biphasic glial-sarcomatous architecture with remnants of the original ependymoma now displaying hypercellularity and atypical - yet not frankly anaplastic - features. The sarcomatous moiety consisted of spindle and epithelioid cells densely interwoven with reticulin fibers. While the ependymal component was GFAP and S100 protein positive, and featured punctate staining for EMA, none of these markers was expressed in the adjacent sarcoma. Instead, the latter reacted for vimentin and smooth muscle actin. To the best of our knowledge, this is the first documentation of tanycytic ependymoma undergoing malignant transformation, one driven by a highly anaplastic mesenchymal component, corresponding to "ependymosarcoma".

Drug hypersensitivity reactions involving skin

Immune reactions to drugs can cause a variety of diseases involving the skin, liver, kidney, lungs, and other organs. Beside immediate, IgE-mediated reactions of varying degrees (urticaria to anaphylactic shock), many drug hypersensitivity reactions appear delayed, namely hours to days after starting drug treatment, showing a variety of clinical manifestations from solely skin involvement to fulminant systemic diseases which may be fatal. Immunohistochemical and functional studies of drug-specific T cells in patients with delayed reactions confirmed a predominant role for T cells in the onset and maintenance of immune-mediated delayed drug hypersensitivity reactions (type IV reactions). In these reactions, drug-specific CD4+ and CD8+ T cells are stimulated by drugs through their T cell receptors (TCR). Drugs can stimulate T cells in two ways: they can act as haptens and bind covalently to larger protein structures (hapten-carrier model), inducing a specific immune response. In addition, they may accidentally bind in a labile, noncovalent way to a particular TCR of the whole TCR repertoire and possibly also major histocompatibility complex (MHC)-molecules - similar to their pharmacologic action. This seems to be sufficient to reactivate certain, probably in vivo preactivated T cells, if an additional interaction of the drug-stimulated TCR with MHC molecules occurs. The mechanism was named pharmacological interaction of a drug with (immune) receptor and thus termed the p-i concept. This new concept may explain the frequent skin symptoms in drug hypersensitivity to oral or parenteral drugs. Furthermore, the various clinical manifestations of T cell-mediated drug hypersensitivity may be explained by distinct T cell functions leading to different clinical phenotypes. These data allowed a subclassification of the delayed hypersensitivity reactions (type IV) into T cell reactions which, by releasing certain cytokines and chemokines, preferentially activate and recruit monocytes (type IVa), eosinophils (type IVb), or neutrophils (type IVd).

Aesthetic outcome and oncological safety of nipple-areola complex replantation after mastectomy and immediate breast reconstruction

Immediate breast reconstruction (IBR) has become an established procedure for women necessitating mastectomy. Traditionally, the nipple-areola complex (NAC) is resected during this procedure. The NAC, in turn, is a principal factor determining aesthetic outcome after breast reconstruction, and due to its particular texture and shape, a natural-looking NAC can barely be reconstructed with other tissues. The aim of this study was to assess the oncological safety as well as morbidity and aesthetic outcome after replantation of the NAC some days after IBR. Retrospective analysis of 85 patients receiving 88 mastectomies and IBR between 1998 and 2007 was conducted. NAC (n=29) or the nipple alone (n=23) were replanted 7 days (median, range 2-10 days) after IBR in 49 patients, provided the subareolar tissue was histologically negative for tumour infiltration. Local recurrence rate was assessed after 49 months (median, range 6-120 months). Aesthetic outcome was evaluated by clinical assessment during routine follow-up at least 12 months after the last intervention. Malignant involvement of the subareolar tissue was found in eight cases (9.1%). Patients qualifying for NAC replantation were in stage 0 in 29%, stage I in 15%, stage IIa in 31%, stage IIb in 17% and stage III in 8%. Total or partial necrosis occurred in 69% and 26% if the entire NAC or only the nipple were replanted, respectively (P<0.01). Depigmentation was seen in 52% and corrective surgery was done in 11 out of 52 NAC or nipple replantations. Local recurrence and isolated regional lymph node metastasis were observed in one single case each. Another 5.8% of the patients showed distant metastases. We conclude that the replantation of the NAC in IBR is oncologically safe, provided the subareolar tissue is free of tumour. However, the long-term aesthetic outcome of NAC replantation is not satisfying, which advocates replanting the nipple alone.

Sciatic neuropathy after body contouring surgery in massive weight loss patients

To date, obesity affects a substantial population in industrialised countries. Due to the increased awareness of obesity-related morbidity, efficient dietary regimens and the recent successes with bariatric surgery, there is now a high demand for body contouring surgery to correct skin abundancies after massive weight loss. The known risks for this type of surgery are mainly wound-healing complications, and, more rarely, thromboembolic or respiratory complications. We present two female patients (23 and 39 years of age) who, in spite of standard positioning and precautions, developed sciatic neuropathy after combined body contouring procedures, including abdominoplasty and inner thigh lift. Complete functional loss of the sciatic nerve was found by clinical and electroneurographic examination on the left side in patient one and bilaterally in patient two. Full nerve conductance recovery was obtained after 6 months in both patients. Although the occurrence of spontaneous neuropathies after heavy weight loss is well documented, this is the first report describing the appearance of such a phenomenon following body contouring surgery. One theoretical explanation may be the compression of the nerve during the semirecumbent positioning combined with hip flexion and abduction, which was required for abdominal closure and simultaneous access to the inner thighs. We advise to avoid this positioning and to include the risk of sciatic neuropathy in the routine preoperative information of patients scheduled for body contouring surgery after heavy weight loss.

The choice of anesthesia influences oxidative energy metabolism and tissue survival in critically ischemic murine skin

In surgical animal studies anesthesia is used regularly. Several reports in the literature demonstrate respiratory and cardiovascular side effects of anesthesiologic agents. The aim of this study was to compare two frequently used anesthesia cocktails (ketamine/xylazine [KX] versus medetomidine/climazolam/fentanyl [MCF]) in skin flap mouse models. Systemic blood values, local metabolic parameters, and surgical outcome should be analyzed in critical ischemic skin flap models. Systemic hypoxia was found in the animals undergoing KX anesthesia compared with normoxia in the MCF group (sO(2): 89.2% +/- 2.4% versus 98.5% +/- 1.2%, P < 0.01). Analysis of tissue metabolism revealed impaired anaerobic oxygen metabolism and increased cellular damage in critical ischemic flap tissue under KX anesthesia (lactate/pyruvate ratio: KX 349.86 +/- 282.38 versus MCF 64.53 +/- 18.63; P < 0.01 and glycerol: KX 333.50 +/- 83.91 micromol/L versus MCF 195.83 +/- 29.49 micromol/L; P < 0.01). After 6 d, different rates of flap tissue necrosis could be detected (MCF 57% +/- 6% versus KX 68% +/- 6%, P < 0.01). In summary we want to point out that the type of anesthesia, the animal model and the goal of the study have to be well correlated. Comparing the effects of KX and MCF anesthesia in mice on surgical outcome was a novel aspect of our study.