Extra-mandibular Osteoradionecrosis after the Treatment of Head and Neck Cancer.

AIMS: Osteoradionecrosis (ORN) is a serious toxicity of head and neck radiotherapy. It predominantly affects the mandible. Extra-mandibular ORN is rare. The aim of this study was to report the incidence and outcomes of extra-mandibular ORNs from a large institutional database. MATERIALS AND METHODS: In total, 2303 head and neck cancer patients were treated with radical or adjuvant radiotherapy. Of these, extra-mandibular ORN developed in 13 patients (0.5%). RESULTS: Maxillary ORNs (n = 8) were a consequence of the treatment of various primaries (oropharynx = 3, sinonasal = 2, maxilla = 2, parotid = 1). The median interval from the end of radiotherapy to the development of ORN was 7.5 months (range 3-42 months). The median radiotherapy dose in the centre of the ORN was 48.5 Gy (range 22-66.5 Gy). Four patients (50%) healed in 7, 14, 20 and 41 months. All temporal bone ORNs (n = 5) developed after treatment to the parotid gland (of a total of 115 patients who received radiotherapy for parotid gland malignancy). The median interval from the end of radiotherapy to the development of ORN was 41 months (range 20-68 months). The median total dose in the centre of the ORN was 63.5 Gy (range 60.2-65.3 Gy). ORN healed in only one patient after 32 months of treatment with repeated debridement and topical betamethasone cream. CONCLUSION: Extra-mandibular ORN is a rare late toxicity and this current study provides useful information on its incidence and outcome. The risk of temporal bone ORN should be considered in the treatment of parotid malignancies and patients should be counselled. More research is required to determine the optimal management of extra-mandibular ORN, particularly on the role of the PENTOCLO regimen.

Iatrogenic T-Cell Lymphoma with Associated Hemophagocytic Lymphohistiocyotsis in a Patient with Long-Standing Rheumatoid Arthritis.

Patients with rheumatoid arthritis are at increased risk of hematological malignancies, especially when exposed to immunosuppressive therapy. The mechanisms of lymphomagenesis remain poorly understood but factors implicated include high disease activity, exposure to antitumoral necrosis factor medications, and Epstein-Barr virus infection. Lymphoid malignancies of T-cell origin are uncommon in patients with rheumatoid arthirits. Clinical presentation with associated hemophagocytic lymphohistiocyotsis is rare and confers a poor prognosis. This case report illustrates a case of a patient with long-standing rheumatoid arthritis and an iatrogenic peripheral T-cell lymphoma with secondary hemophagocytic lymphohistiocytosis who achieved a complete response after intensive chemotherapy.

Perinucleolar compartment prevalence has an independent prognostic value for breast cancer.

The perinucleolar compartment (PNC) is a multicomponent nuclear structure enriched with RNAs transcribed by RNA pol III and RNA binding proteins. Studies in cultured cells showed an association between PNC and transformed phenotype. To evaluate the relationship between structure and malignancy in vivo, we examined PNC prevalence (the percentage of cells containing at least one PNC) in normal and cancerous paraffin-embedded breast tissues using immunohistochemistry against a PNC-associated protein. Five hundred nuclei in the most active area of each sample were scored for PNC prevalence. The results show that PNC prevalence significantly correlates with the progression of breast cancer (by the criteria of staging). PNC prevalence in primary tumors, lymph nodes, and distant metastases shows a stepwise increase from a median of 23% in primary tumors to approximately 100% in distant metastases. In addition, univariate and multivariate (controlling for tumor size and grade) analyses show that early-stage patients with invasive ductal carcinomas containing a higher PNC prevalence have a significantly poorer prognosis. These findings link PNC prevalence with the progression of breast cancer in vivo and suggest that PNC-containing cells have metastatic advantages. These findings also show the potential of PNC prevalence as a prognostic marker for breast cancer.

Histological characterization of the thyroglossal tract: implications for surgical management.

OBJECTIVES: In the current report, we characterized the relationship between the central hyoid bone and the thyroglossal tract and determined the prevalence of ectopic thyroid follicles in the adjacent soft tissues. STUDY DESIGN: Retrospective pathological analysis. METHODS: The resected specimens from 104 patients who underwent a modified Sistrunk procedure with wide-field dissection were retrospectively analyzed. Under light microscopy, serial sections were examined to determine whether the thyroglossal tract passed anterior to, posterior to, or within the hyoid bone. Specimens were also examined for the presence of thyroid follicles in the periductal and pericystic soft tissues. RESULTS: In 50 cases (48%), the tract position in relation to the hyoid bone could not be identified secondary to extensive arborization, marked inflammation, specimen fragmentation or a combination of these. Thyroid follicles were observed in 9 (18%) of these specimens. In the remaining 54 cases (52%) the tract was located anterior to the central arch of the hyoid bone in 39 (72%) and posterior to it in 15 (28%). Thyroid tissue was observed in 46% of specimens (P =.004), regardless of whether the tract was anterior or posterior. CONCLUSIONS: These results demonstrate that tract position often cannot be defined, but when a portion is dominant, it is likely to be anterior to the hyoid bone. Ectopic thyroid tissue can be found in almost 50% of specimens when the tract position is identifiable. With appropriate surgical management, a recurrence rate of less than 4% can be expected, despite the presence of ductule multiplicity, marked inflammation, tract position posterior to the hyoid bone, and ectopic thyroid follicles.

Analysis of tamoxifen and its metabolites in synthetic gastric fluid digests and urine samples using high-performance liquid chromatography with electrospray mass spectrometry.

We report on the transformation of tamoxifen at 37 degrees C in synthetic gastric fluid as studied by high-performance liquid chromatography with triple quadrupole mass spectrometry. The major transformation products detected were (E)-isomer of tamoxifen, metabolite D, and several unidentified components having m/z 404. Addition of pepsin to the gastric fluid inhibited formation of all of these products. We analyzed several urine samples from breast cancer patients undergoing tamoxifen treatment. Metabolite D was identified in the urine samples and in the gastric fluid digest at a retention time of 22.0 min eluting from a reversed-phase HPLC column. Although several metabolites were found in all the urine samples of patients, some metabolites were detected in one sample but not others, suggesting tamoxifen metabolism varies in patients.

Detection of EBV DNA in the cord blood donor for a patient developing Epstein-Barr virus-associated lymphoproliferative disorder following mismatched unrelated umbilical cord blood transplantation.

Epstein-Barr virus-associated post-transplant lymphoproliferative disorder (PTLD) has been well described as a complication following allogeneic stem cell transplantation but has only recently been reported following umbilical cord blood (UCB) transplant. We report the case of a child transplanted with unrelated mismatched UCB for juvenile chronic myelogenous leukemia (JCML) who developed EBV-associated PTLD, which was confirmed pathologically, 139 days following stem cell infusion. There was no clinical response to reduction of immune suppression, high-dose acyclovir, or alpha interferon. The patient died 160 days after transplantation. EBV was detected by polymerase chain reaction in the cord blood unit used for transplantation. This case demonstrates that EBV-associated PTLD can occur following mismatched unrelated UCB transplant and may be related to transmission of EBV infection by donor lymphocytes.

Autoimmune hepatitis as a late complication of liver transplantation.

BACKGROUND: The development of de novo autoimmune hepatitis as a long-term complication after liver transplantation has been recently reported. The authors describe five liver allograft recipients who developed chronic hepatitis associated with autoimmune features. METHODS: Five of 155 liver transplant recipients at risk (2.5%) developed this particular form of graft dysfunction. The authors review the clinical records, liver histology, therapy, and outcome of these five patients. RESULTS: Patients included two boys and three girls. Median age at transplantation was 3.5 years (range, 0.5-14 years), median age at presentation was 9 years (range, 2-17 years), and median interval after transplantation was 5.1 years (range, 1.5-9 years). Indications for liver transplant included biliary atresia in four patients and primary sclerosing cholangitis in one patient. At the time of presentation, all patients were receiving cyclosporine as their primary immunosuppressive agent. Only one patient had a history of rejection, which had resolved. All patients presented with increased transaminase levels, and one had a mildly elevated conjugated bilirubin level. Only one patient had constitutional complaints. Acute and chronic rejection, viral hepatitis, vascular insufficiency, and biliary tract obstruction were excluded. Antinuclear antibody levels were elevated in four patients (titer range, 1:160-1:640), one of whom also had positive antismooth muscle antibody (titer 1:80) results. The fifth patient had an elevated serum total protein level. Histologic analysis of liver biopsy samples from the five patients showed findings consistent with chronic autoimmune hepatitis. All patients were treated with standard therapy for autoimmune hepatitis, which included daily steroids and azathioprine. Cyclosporine doses were reduced in three patients and eliminated in two. All patients responded with normalization (n = 2) or improvement (n = 3) of liver transaminases within the first 3 months of therapy. Histologic analysis of the 3-month follow-up liver biopsy was normal (n = 2) or showed improvement in inflammation (n = 2). Two patients developed acute allograft rejection within 6 to 12 months after discontinuation or reduction in cyclosporine. CONCLUSIONS: Autoimmune hepatitis occurs after liver transplantation in patients without a previous history of autoimmune hepatitis. The risk of developing autoimmune hepatitis appears to be greater in children after liver transplantation than in the general pediatric population. Standard therapy for autoimmune hepatitis is effective.

Natural killer cell lymphoma: report of two pediatric cases, therapeutic options, and review of the literature.

BACKGROUND: Natural killer (NK) cell lymphomas are rapidly fatal malignancies that to the authors' knowledge are rare in children. In the current study, the authors report the cases of two boys with NK cell lymphomas with refractory disease who both were salvaged with high dose chemotherapy and stem cell transplantation and compare these patients with those in the published experience. METHODS: A comprehensive literature review was performed to identify other cases of pediatric patients with NK cell lymphomas, their treatment, and outcome. RESULTS: One of the patients in the current study developed two recurrences and the other patient experienced early disease progression during front-line treatment. Both then were treated with high dose chemotherapy followed by stem cell rescue. At last follow-up, the patients remained free of disease at 15 months and 16 months, respectively, after transplantation (48 months and 22 months, respectively, from the time of diagnosis). In addition to the 2 patients in the current study, the authors found 13 pediatric patients reported in the literature to date. Of the 7 patients with localized (Stage I-II) disease, 5 patients (71%) were reported to be alive 1-107 months after diagnosis. Of the 6 patients with Stage IV disease, only the 2 patients who received high dose chemotherapy and stem cell rescue (33%) were alive at the time of last follow-up (at 30 months and 12 months, respectively). Including the patients reported in the current study, 9 of 15 children with NK cell lymphoma (all stages) (60%) were reported to be alive at the time of last follow-up. CONCLUSIONS: Although pediatric NK cell lymphomas rapidly can become fatal, it appears that high dose chemotherapy followed by stem cell transplantation is effective therapy, especially in patients with advanced or resistant disease. Further follow-up is needed to determine whether this treatment approach will be curative.

Specificity of signaling by STAT1 depends on SH2 and C-terminal domains that regulate Ser727 phosphorylation, differentially affecting specific target gene expression.

Complete activation of signal transducer and activator of transcription 1 (STAT1) requires phosphorylation at both Y701 and a conserved PMS(727)P sequence. S727 phosphorylation of STAT1 in interferon-gamma (IFN-gamma)-treated mouse fibroblasts occurred without a need for p38 mitogen-activated protein kinase (MAPK), extracellular signal-regulated kinases 1 and 2 or c-Jun kinases, and required both an intact SH2 domain and phosphorylation of Y701. In contrast, UV irradiation-induced STAT1 phosphorylation on S727 required p38MAPK, but no SH2 domain- phosphotyrosine interactions. Mutation of S727 differentially affected IFN-gamma target genes, at the level of both basal and induced expression. Particularly strong effects were noted for the GBP1 and TAP1 genes. The PMS(727)P motif of STAT3 was phosphorylated by stimuli and signaling pathways different from those for STAT1 S727. Transfer of the STAT3 C-terminus to STAT1 changed the stimulus and pathway specificity of STAT1 S727 phosphorylation to that of STAT3. Our data suggest that STAT C-termini contribute to the specificity of cellular responses by linking individual STATs to different serine kinase pathways and through an intrinsically different requirement for serine phosphorylation at different target gene promoters.

Partial impairment of cytokine responses in Tyk2-deficient mice.

To assess the role of the Janus kinase (Jak) family member Tyk2, we have generated Tyk2-/- mice. In contrast to other Jaks, where inactivation leads to a complete loss of the respective cytokine receptor signal, Tyk2-/- mice display reduced responses to IFNalpha/beta and IL-12 and a selective deficiency in Stat3 activation in these pathways. Unexpectedly, IFNgamma signaling is also impaired in Tyk2-/- mice. Tyk2-/- macrophages fail to produce nitric oxide upon lipopolysaccharide induction. Tyk2-/- mice are unable to clear vaccinia virus and show a reduced T cell response after LCMV challenge. These data imply a selective contribution of Tyk2 to the signals triggered by various biological stimuli and cytokine receptors.

Serine phosphorylation of STATs.

Tyrosine phosphorylation regulates the dimerization of STATs as an essential prerequisite for the establishment of a classical JAK-STAT signaling path. However, most vertebrate STATs contain a second phosphorylation site within their C-termini. The phosphorylated residue in this case is a serine contained within a P(M)SP motif, and in the majority of situations its mutation to alanine alters transcription factor activity. This review addresses recent advances in understanding the regulation of STAT serine phosphorylation, as well as the kinases and other signal transducers implied in this process. The biochemical and biological consequences of STAT serine phosphorylation are discussed. Oncogene (2000).

Rpg1p, the subunit of the Saccharomyces cerevisiae eIF3 core complex, is a microtubule-interacting protein.

The essential gene RPG1/TIF32 of Saccharomyces cerevisiae encodes the 110-kDa subunit of the translation initiation factor 3 (eIF3) core complex. In this study, the Rpg1p-specific monoclonal antibody PK1/1 was used to analyse the cellular distribution of Rpg1p by epifluorescence and confocal laser scanning microscopy (CLSM). In budded cells, a portion of Rpg1p was obviously co-localised with microtubules. In addition, CLSM revealed an accumulation of Rpg1p in a patch at the very end of cytoplasmic microtubules reaching the bud tip. A punctate fluorescence pattern was typical for separated unbudded cells. Distribution of Rpg1p was confirmed using a strain expressing exclusively a hemaglutinin-tagged version of Rpg1p. In nocodazole-treated cells, the pattern of the PK1/1 staining was disturbed. No staining was observed in Rpg1p-depleted cells. In vitro experiments revealed that Rpg1p was specifically co-immunoprecipitated with alpha-tubulin from the yeast cell free extract and this observation was further supported by showing that Rpg1p co-sedimented with hog brain microtubules. We conclude that Rpg1p is a microtubule-interacting protein that indicates an interesting connection between the translation initiation machinery and cytoskeleton in yeast Saccharomyces cerevisiae.

Stress-induced phosphorylation of STAT1 at Ser727 requires p38 mitogen-activated protein kinase whereas IFN-gamma uses a different signaling pathway.

STAT1 is an essential transcription factor for macrophage activation by IFN-gamma and requires phosphorylation of the C-terminal Ser727 for transcriptional activity. In macrophages, Ser727 phosphorylation in response to bacterial lipopolysaccharide (LPS), UV irradiation, or TNF-alpha occurred through a signaling path sensitive to the p38 mitogen-activated protein kinase (p38 MAPK) inhibitor SB203580 whereas IFN-gamma-mediated Ser727 phosphorylation was not inhibited by the drug. Consistently, SB203580 did not affect IFN-gamma-mediated, Stat1-dependent transcription but inhibited its enhancement by LPS. Furthermore, LPS, UV irradiation, and TNF-alpha caused activation of p38 MAPK whereas IFN-gamma did not. An essential role for p38 MAPK activity in STAT1 Ser727 phosphorylation was confirmed by using cells expressing an SB203580-resistant p38 MAPK. In such cells, STAT1 Ser727 phosphorylation in response to UV irradiation was found to be SB203580 insensitive. Targeted disruption of the mapkap-k2 gene, encoding a kinase downstream of p38 MAPK with a key role in LPS-stimulated TNF-alpha production and stress-induced heat shock protein 25 phosphorylation, was without a significant effect on UV-mediated Ser727 phosphorylation. The recombinant Stat1 C terminus was phosphorylated in vitro by p38MAPKalpha and beta but not by MAPK-activated protein kinase 2. Janus kinase 2 activity, previously reported to be required for IFN-gamma-mediated Ser727 phosphorylation, was not needed for LPS-mediated Ser727 phosphorylation, and activation of Janus kinase 2 did not cause the appearance of STAT1 Ser727 kinase activity. Our data suggest that STAT1 is phosphorylated at Ser727 by a stress-activated signaling pathway either through p38 MAPK directly or through an unidentified kinase downstream of p38MAPK.

Transcription factor activity of STAT proteins: structural requirements and regulation by phosphorylation and interacting proteins.

The seven mammalian members of the signal transducer and activator of transcription (STAT) family share a common core structure which reflects their shared mechanism of activation, dimerization, and DNA binding. By contrast, the STAT C termini containing the sequences required for transcriptional activation are much less homologous, suggesting different ways by which individual STATs activate their target genes. This paper describes several important discoveries linked to mechanistic aspects of STAT transcription factor function. These include regulated serine phosphorylation of the transactivating domain, promoter-dependent interactions of STATs with each other, or of STATs with other transcription factors, and with transcriptional co-activators. The basis, background, and implications of these molecular events will be summarized and discussed.

Lipopolysaccharide induces in macrophages the synthesis of the suppressor of cytokine signaling 3 and suppresses signal transduction in response to the activating factor IFN-gamma.

The goal of this study was to investigate how bacterial LPS affects macrophage responsiveness to the activating factor IFN-gamma. Pretreatment of macrophages with LPS for <2 h increased the transcriptional response to IFN-gamma. In contrast, simultaneous stimulation with IFN-gamma and LPS, or pretreatment with LPS for >4 h, suppressed Stat1 tyrosine 701 phosphorylation, dimerization, and transcriptional activity in response to IFN-gamma. Consistently, the induction of MHCII protein by IFN-gamma was antagonized by LPS pretreatment. Neutralizing Abs to IL-10 were without effect on LPS-mediated suppression of Stat1 activation. Decreased IFN-gamma signal transduction after LPS treatment corresponded to a direct induction of suppressor of cytokine signaling3 (SOCS3) mRNA and protein. Under the same conditions socs1, socs2, and cis genes were not transcribed. In transfection assays, SOCS3 was found to suppress the transcriptional response of macrophages to IFN-gamma. A causal link of decreased IFN-gamma signaling to SOCS3 induction was also suggested by the LPS-dependent reduction of IFN-gamma-mediated Janus kinase 1 (JAK1) activation. Further consistent with inhibitory activity of SOCS3, LPS also inhibited the JAK2-dependent activation of Stat5 by GM-CSF. Our results thus link the deactivating effect of chronic LPS exposure on macrophages with its ability to induce SOCS3.

Protein tyrosine kinase Pyk2 mediates the Jak-dependent activation of MAPK and Stat1 in IFN-gamma, but not IFN-alpha, signaling.

Two distinct types of interferon, IFN-alpha/beta and IFN-gamma, commonly exhibit antiviral activities by transmitting signals to the interior of the cell via their homologous receptors. Receptor stimulation results in the activation of distinct combinations of Janus family protein tyrosine kinases (Jak PTKs); Jak1/Tyk2 and Jak1/Jak2 for IFN-alpha/beta and IFN-gamma, respectively. Jak PTK activation by these IFNs is commonly followed by tyrosine phosphorylation of the transcription factor Stat1 at Y701, which is essential for dimerization, translocation to the nucleus and DNA-binding activity. To gain full transcriptional activity, Stat1 also requires serine phosphorylation at S727. In this paper we demonstrate that Pyk2, which belongs to another PTK family, is critical for the Jak-mediated MAPK and Stat1 activation by IFN-gamma, but not IFN-alpha. Pyk2 is selectively associated with Jak2 and activated by IFN-gamma. Overexpression of PKM, a dominant interfering form of Pyk2, in NIH 3T3 cells results in a strong inhibition of the IFN-gamma-induced activation of Erk2, serine phosphorylation of Stat1 and Stat1-dependent gene transcription. Finally, the antiviral action of IFN-gamma, but not IFN-alpha, is severely impaired by PKM overexpression. Thus, the two types of IFN may utilize distinct Jak-mediated Erk2, and possibly other MAPK activation pathways for their antiviral action.

Salmonella typhimurium and lipopolysaccharide stimulate extracellularly regulated kinase activation in macrophages by a mechanism involving phosphatidylinositol 3-kinase and phospholipase D as novel intermediates.

Activation of the extracellularly regulated kinase (ERK) pathway is part of the early biochemical events that follow lipopolysaccharide (LPS) treatment of macrophages or their infection by virulent and attenuated Salmonella strains. Phagocytosis as well as the secretion of invasion-associated proteins is dispensable for ERK activation by the pathogen. Furthermore, the pathways used by Salmonella and LPS to stimulate ERK are identical, suggesting that kinase activation might be solely mediated by LPS. Both stimuli activate ERK by a mechanism involving herbimycin-dependent tyrosine kinase(s) and phosphatidylinositol 3-kinase. Phospholipase D activation and stimulation of protein kinase C appear to be intermediates in this novel pathway of MEK/ERK activation.

Gastric spiral bacteria in small felids.

Nine small cats, including one bobcat (Felis rufus), one Pallas cat (F. manul), one Canada lynx (F. lynx canadensis), two fishing cats (F. viverrina), two margays (F. wiedii), and two sand cats (F. margarita), necropsied between June 1995 and March 1997 had large numbers of gastric spiral bacteria, whereas five large cats, including one African lion (Panthera leo), two snow leopards (P. uncia), one Siberian tiger (P. tigris altaica), and one jaguar (P. onca), necropsied during the same period had none. All of the spiral organisms from the nine small cats were histologically and ultrastructurally similar. Histologically, the spiral bacteria were 5-14 microm long with five to nine coils per organism and were located both extracellularly within gastric glands and surface mucus, and intracellularly in parietal cells. Spiral bacteria in gastric mucosal scrapings from the Canada lynx, one fishing cat, and the two sand cats were gram negative and had corkscrewlike to tumbling motility when viewed with phase contrast microscopy. The bacteria were 0.5-0.7 microm wide, with a periodicity of 0.65-1.1 microm in all cats. Bipolar sheathed flagella were occasionally observed, and no periplasmic fibrils were seen. The bacteria were extracellular in parietal cell canaliculi and intracellular within parietal cells. Culture of mucosal scrapings from the Canada lynx and sand cats was unsuccessful. Based on morphology, motility, and cellular tropism, the bacteria were probably Helicobacter-like organisms. Although the two margays had moderate lymphoplasmacytic gastritis, the other cats lacked or had only mild gastric lymphoid infiltrates, suggesting that these organisms are either commensals or opportunistic pathogens.