Indeterminate nodules in osteosarcoma: what's the follow-up?

BACKGROUND: Indeterminate pulmonary nodules in patients diagnosed with osteosarcoma present a challenge for accurate staging and prognosis. The aim of this study was to explore the significance of this finding. METHODS: A retrospective cohort study of 120 patients with osteosarcoma was performed in the North East of England. Chest computed tomographies (CTs) at presentation were reviewed and the incidence of 'indeterminate' nodules recorded. Follow-up scans were reviewed and survival as well as prognostic features were analysed. RESULTS: 25% of our cohort presented with indeterminate nodules. Of these, 33% were subsequently confirmed as metastases, the majority within a year. Kaplan-Meier survival analysis showed that patients with indeterminate nodules fared better than those with frank metastatic disease, and similar to those who presented with a normal chest CT. We found no radiographic features that predicted survival. CONCLUSIONS: Indeterminate nodules remain a clinical and diagnostic dilemma. Close monitoring of patients is advised during the first year from presentation, and there is potential for indeterminate nodules to develop into frank metastases later than five years from presentation.

Ileocaecal junction carcinoma: a clinicopathological study of 199 cases.

AIM: The ileocaecal junction (ICJ) region is an epithelial transition zone in which carcinomas are frequently diagnosed. However, it is currently unknown whether ICJ carcinomas (ICJ-CAs) have distinctive features. This study aimed to characterize the clinicopathological features of ICJ-CAs. METHOD: All ileal and colorectal resections for carcinoma, performed in Calgary, Canada between January 2009 and June 2012, were reviewed. Carcinomas in which the epicentre was within 5 cm of the ileocaecal valve (ICV) were defined as ICJ-CAs. Of 1003 carcinomas studied, 199 (19.8%) were ICJ-CAs, including 93 (9.3%) that crossed the ICV. Comparison of clinicopathological features with carcinomas of the other ileo-colorectal regions was made. Survival was also assessed. RESULTS: Clinically, ICJ-CAs were more common in female than male patients (56.3% female) compared with left-colonic (42.9% female) and rectal (37.9% female) carcinomas, and were more common in older age-groups of patients (71.8 ± 12.7 years) compared with appendiceal (62.6 ± 11.3 years), left-colonic (69.4 ± 12.3 years) and rectal (67.1 ± 11.9 years) carcinomas. Macroscopically, ICJ-CAs were similar to other colorectal carcinomas and were mostly described as ulcerated (63.3%). Histologically, ICJ-CAs had more mucinous, signet-ring cell and/or neuroendocrine features (39.7%, 8.0% and 7.5%, respectively) than did carcinomas of the left colon (16.8%, 1.6% and 1.1%, respectively) and the rectum (14.1%, 1.0% and 0.0%, respectively). They were higher grade (20.1% were high grade) than those of the left-colon (10.3%) and the rectum (9.8%). ICJ-CAs presented at a higher T-stage (25.6% were T4) compared with rectal carcinomas (11.6%). Most significantly, ICJ-CAs presented at a higher N-stage (25.6% were N2) than did right-colonic (14.1%) and rectal (16.2%) carcinomas. Although survival of patients with ICJ-CAs did not differ from those with right-colonic carcinomas, those with carcinomas directly involving the ICV did show a significantly decreased survival. CONCLUSION: ICJ-CAs display several distinct clinicopathological features that may require special diagnostic, prognostic and management attention.

Analysis of Antarctic proteobacteria by PCR fingerprinting and screening for antimicrobial secondary metabolites.

Fifty-seven proteobacterium species were successfully isolated from soils of Barrientos Island of the Antarctic using 11 different isolation media. Analysis of 16S rDNA sequencing of these isolates showed that they belonged to eight different genera, namely Bradyrhizobium, Sphingomonas, Methylobacterium, Caulobacter, Paracoccus, Ralstonia, Rhizobium, and Staphylococcus. All isolates were studied for capability of producing antimicrobial and antifungal secondary metabolites using high-throughput screening models. Approximately 23 (13/57) and 2% (1/57) of isolates inhibited growth of Candida albicans ATCC 10231(T) and Staphylococcus aureus ATCC 51650(T), respectively. These results indicated that proteobacterium species isolates from Antarctic could serve as potential source of useful bioactive metabolites. Enterobacterial repetitive intergenic consensus (ERIC)-PCR fingerprinting produced nine clusters and 13 single isolates, with a high D value of 0.9248. RAPD fingerprinting produced six clusters and 13 single isolates, with a relatively low D value of 0.7776. ERIC-PCR analysis proved to have better discrimination capability than RAPD analysis and generated better clustering for all proteobacterium species isolates. We conclude that ERIC-PCR is a robust, reliable and rapid molecular typing method for discriminating different genera of proteobacteria.

Rescue therapy with local intra-arterial urokinase after poor clinical response with intravenous recombinant tissue plasminogen activator in acute ischaemic stroke.

We report a case of combined intravenous and intra-arterial thrombolysis in a patient presenting with acute ischaemic stroke. Progression to intra-arterial thrombolysis in patients who fail to show clinical improvement within 6 h of symptom onset might be a safe treatment option at centres with access to stroke specialist and endovascular services. The purpose of this report is to demonstrate the efficacy and potential benefits of this rescue therapy.

An abnormal nonhyperdiploid karyotype is a significant adverse prognostic factor for multiple myeloma in the bortezomib era.

Multiple myeloma is clinically heterogeneous and risk stratification is vital for prognostication and informing treatment decisions. As bortezomib is able to overcome several high-risk features of myeloma, the validity of conventional risk-stratification and prognostication systems needs to be reevaluated. We study the survival data of 261 previously untreated myeloma patients managed at our institution, where bortezomib became available from 2004 for the treatment of relapse disease. Patient and disease characteristics, and survival data were evaluated overall, and with respect to bortezomib exposure. Overall, the international staging system (ISS), metaphase karyotyping and interphase fluorescence in situ hybridization (FISH) were discerning of survival outcomes, where the median for the entire cohort was 5.2 years. However, when stratified by bortezomib exposure, only metaphase karyotyping was still discriminating of long-term prognosis. The presence of an abnormal nonhyperdiploid karyotype overrides all other clinical and laboratory parameters in predicting for a worse outcome on multivariate analysis (median survival 2.6 years, P = 0.001), suggesting that bortezomib used at relapse is better able to overcome adverse risk related to high tumor burden (as measured by the ISS) than adverse cytogenetics on conventional karyotyping. Metaphase karyotyping provides additional prognostic information on tumor kinetics where the presence of a normal diploid karyotype in the absence of any high-risk FISH markers correlated with superior survival and could act as a surrogate for lower plasma cell proliferation.

Acquired factor V deficiency in a patient with myeloma and amyloidosis.

INTRODUCTION: We describe our experience with managing an unusual case of acquired Factor V deficiency (aFVd) in a myeloma patient with demonstrated amyloidosis. METHODS: Following diagnosis, records of previous investigations were sought. Specific clotting factors and inhibitors were tested. The clinical progress and treatment response measured by serial factor V levels and coagulation parameters was then prospectively tracked. RESULTS: A 57 year-old woman presented with spontaneous right knee haemarthrosis in association with bilateral symmetrical polyneuropathy and proteinuria. Coagulation screen showed prolongation of both PT (18.6 s, normal range [9.9-11.4 s]) and aPTT (41.4 s, normal range [25.7-32.9 s]), which were both fully correctable following a mixing study. Liver function, fibrinogen, clotting factor II/VIII/X assays and disseminated intravascular coagulopathy screen was normal. FV level was reduced (19%, normal range [70-170%]). Inhibitor titer was undetectable. Congenital FVd was excluded as her previous coagulation screen was normal. Bone marrow investigation performed for suspected underlying plasma cell dyscrasia showed 60% neoplastic plasma cells. Congo red staining was positive for amyloid within vascular walls of the marrow trephine. She was diagnosed with light chain myeloma and aFVd. She received Bortezomib/Cyclophosphamide/Dexamethasone (VCD) chemotherapy. After one cycle of VCD, serum kappa free light chain (SFLC) was reduced from 6951 mg/L to 3354 mg/L with serial measurements of FV levels showing increment to 76% and normalization of PT/aPTT. CONCLUSION: Plasma cell dyscrasia with amyloidosis should be sought as a cause for aFVD, in particular one where bleeding manifestation is profound even with the absence of demonstrable inhibitors.

Cytoplasmic sequestration of rel proteins by IkappaBalpha requires CRM1-dependent nuclear export.

Rel and IkappaB protein families form a complex cellular regulatory network. A major regulatory function of IkappaB proteins is to retain Rel proteins in the cell cytoplasm. In addition, IkappaB proteins have also been postulated to serve nuclear functions. These include the maintenance of inducible NF-kappaB-dependent gene transcription, as well as termination of inducible transcription. We show that IkappaBalpha shuttles between the nucleus and the cytoplasm, utilizing the nuclear export receptor CRM1. A CRM1-binding export sequence was identified in the N-terminal domain of IkappaBalpha but not in that of IkappaBbeta or IkappaBepsilon. By reconstituting major aspects of NF-kappaB-IkappaB sequestration in yeast, we demonstrate that cytoplasmic retention of p65 (also called RelA) by IkappaBalpha requires Crm1p-dependent nuclear export. In mammalian cells, inhibition of CRM1 by leptomycin B resulted in nuclear localization of cotransfected p65 and IkappaBalpha in COS cells and enhanced nuclear relocation of endogenous p65 in T cells. These observations suggest that the main function of IkappaBalpha is that of a nuclear export chaperone rather than a cytoplasmic tether. We propose that the nucleus is the major site of p65-IkappaBalpha association, from where these complexes must be exported in order to create the cytoplasmic pool.

Enzymatic formation of prostaglandin D2 by rat basophilic leukemia cells and normal rat mast cells.

It has been shown that the major cyclooxygenase product in rat basophilic leukemia (RBL-1) cells and in normal rat mast cells is prostaglandin D2. In RBL-1 cells, prostaglandin D2 is isomerase activity was found in the 150 000 X g microsomal pellet as well as the supernatant fraction. Incubation of RBL-1 microsomes with arachidonic acid without cofactors yielded 17.5 +/- 2% prostaglandin E2 and 9.1 +/- 1.4% prostaglandin D2. The cyclooxygenase activity was enhanced (25%) by epinephrine and the addition of reduced glutathione led to a marked increase in prostaglandin D2 synthesis (3-fold). Incubations with arachidonic acid, glutathione and epinephrine gave the maximum conversion to prostaglandin D2, yielding 7 +/- 0.4% prostaglandin E2 and 35.6 +/- 3.5% prostaglandin D2. Incubations with [14C]prostaglandin H2 to bypass cyclooxygenase confirmed the presence and glutathione dependence of the prostaglandin D2 isomerase in the microsomal fraction and also revealed the presence of the same enzyme in the 150 000 X g supernant. In contrast to RBL-1 cells, incubations of microsomes and supernatant from normal rat mast cells with [14C]-arachidonic acid and [14C]prostaglandin H2 localized the prostaglandin D2 isomerase activity in the soluble fraction. Similar to the enzyme in the RBL-1 cells, the mast cell enzyme was glutathione dependent.

Deep-vein thrombosis rates after major orthopedic surgery in Asia. An epidemiological study based on postoperative screening with centrally adjudicated bilateral venography.

BACKGROUND: The incidence of postsurgical venous thromboembolism is thought to be low in Asian ethnic populations. OBJECTIVE: We studied the incidence of deep-vein thrombosis (DVT) in Asian patients undergoing major orthopedic surgery of the lower limbs. PATIENTS/METHODS: We performed a prospective epidemiological study in 19 centers across Asia (China, Indonesia, South Korea, Malaysia, Philippines, Taiwan, and Thailand) in patients undergoing elective total hip replacement (THR), total knee replacement (TKR) or hip fracture surgery (HFS) without pharmacological thromboprophylaxis. The primary endpoint was the rate of DVT of the lower limbs documented objectively with bilateral ascending venography performed 6-10 days after surgery using a standardized technique and evaluated by a central adjudication committee unaware of local interpretation. RESULTS: Overall, of 837 Asian patients screened for this survey, 407 (48.6%, aged 20-99 years) undergoing THR (n = 175), TKR (n = 136) or HFS (n = 96) were recruited in 19 centers. DVT was diagnosed in 121 of 295 evaluable patients [41.0%, (95% confidence interval (CI): 35.4-46.7)]. Proximal DVT was found in 30 patients [10.2% (7.0-14.2)]. Total DVT and proximal DVT rates were highest in TKR patients (58.1% and 17.1%, respectively), followed by HFS patients (42.0% and 7.2%, respectively), then THR patients (25.6% and 5.8%, respectively). DVT was more frequent in female patients aged at least 65 years. Pulmonary embolism was clinically suspected in 10 of 407 patients (2.5%) and objectively confirmed in two (0.5%). CONCLUSIONS: The rate of venographic thrombosis in the absence of thromboprophylaxis after major joint surgery in Asian patients is similar to that previously reported in patients in Western countries.

Outcome of carpal tunnel release--correlation with wrist and wrist-palm anthropomorphic measurements.

Wrist and wrist-palm measurements have been associated with the diagnosis of carpal tunnel syndrome. We found no reported study about how this correlation affects the outcome after surgery. We investigated the role of the measurements in predicting outcome after open carpal tunnel release. A total of 131 patients (88 female, 43 male) responded to our postal questionnaire using the Boston Carpal Tunnel assessment (65% response rate) at a minimum of 9 months post-operatively. Symptom and functional scores showed a strong correlation. There was no statistical difference in the outcome between wrist ratio (≥0.7 vs <0.7), wrist-palm ratio (≥0.41 vs <0.41) and gender, but a better functional score was very weakly correlated with a higher wrist ratio. A very large study would be needed to show any statistical correlation between both measurement and outcome.

Efficacy and safety of edoxaban for treatment of venous thromboembolism: a subanalysis of East Asian patients in the Hokusai-VTE trial.

BACKGROUND: Direct oral anticoagulants have been evaluated for their efficacy and safety in the treatment of venous thromboembolism (VTE), which comprises deep vein thrombosis and pulmonary embolism. The randomized, double-blind Hokusai-VTE trial demonstrated that 60 mg of edoxaban once daily following initial heparin treatment is non-inferior to heparin overlapped with and followed by warfarin for the treatment of VTE, and is associated with significantly fewer bleeding events. OBJECTIVES: To assess the efficacy and safety of edoxaban versus warfarin among East Asian patients enrolled in the Hokusai-VTE trial. PATIENTS/METHODS: The Hokusai-VTE trial enrolled 8292 patients from 439 centers worldwide, including 1109 patients from Japan, China, Korea, and Taiwan. The primary efficacy and safety outcomes were symptomatic recurrent VTE and clinically relevant bleeding, respectively. RESULTS: In the overall East Asian population, the primary efficacy outcome of symptomatic recurrent VTE occurred in 16 of 563 (2.8%) patients in the edoxaban group versus 24 of 538 (4.5%) patients in the warfarin group (hazard ratio [HR] 0.64; 95% confidence interval [CI] 0.34-1.19; P = 0.1601). The primary safety outcome of clinically relevant bleeding occurred in 56 of 563 (9.9%) patients in the edoxaban group versus 93 of 538 (17.3%) patients in the warfarin group (HR 0.56; 95% CI 0.40-0.78; P < 0.001). CONCLUSIONS: Edoxaban is an effective and safer alternative to warfarin in East Asian patients with acute VTE who require anticoagulant therapy, consistent with overall study findings from the Hokusai-VTE trial.

Molecular characterization of KEX1, a kexin-like protease in mouse Pneumocystis carinii.

Expression screening of a Pneumocystis carinii-infected mouse lung cDNA library with specific monoclonal antibodies (mAbs) led to the identification of a P. carinii cDNA with extensive homology to subtilisin-like proteases, particularly fungal kexins and mammalian prohormone convertases. The 3.1 kb cDNA contains a single open reading frame encoding 1011 amino acids. Structural similarities to fungal kexins in the deduced primary amino acid sequence include a putative proenzyme domain delineated by a consensus autocatalytic cleavage site (Arg-Glu-Lys-Arg), conserved Asp, His, Asn and Ser residues in the putative catalytic domain, a hydrophobic transmembrane spanning domain, and a carboxy-terminal cytoplasmic domain with a conserved tyrosine motif thought to be important for localization of the protease in the endoplasmic reticulum and/or Golgi apparatus. Based on these structural similarities and the classification of P. carinii as a fungus, the protease was named KEX1. Southern blotting of mouse P. carinii chromosomes localized kex1 to a single chromosome of approximately 610 kb. Southern blotting of restriction enzyme digests of genomic DNA from P. carinii-infected mouse lung demonstrated that kex1 is a single copy gene. The function of kexins in other fungi suggests that KEX1 may be involved in the post-translational processing and maturation of other P. carinii proteins.

Timing and patterns of astrocyte migration from xenogeneic transplants of the cortex and corpus callosum.

The timing, pattern, and pathway of astrocyte migration were investigated in vivo by transplantation of CD-1 mouse cerebral cortex (E13-14) or corpus callosum (P2-3) into neonatal rat cortex. A monoclonal antibody specific for a mouse astrocyte surface antigen (M2) was used to identify the location of the grafts and the migrated donor astrocytes. Within the host cortex, astrocytes from cortical grafts began migration at post-transplantation day (PTD) 7. Over the next 4 days, the most distant displaced donor cells were found progressively further away from the grafts, migrating at a rate of about 220 microns/day. After PTD 11, the migration rate for the farthest displaced donor cells slowed to 25 microns/day, and the cells appeared to stop at about PTD 16 at a distance of 1,100 microns from the edge of the graft. Astrocytes had a faster migration speed in the white matter and covered a longer distance (5 mm) than those in the gray matter, extending on occasion into the contralateral hemisphere. The patterns of astrocyte migration differed depending on local cues around the transplant. Donor astrocytes that had been implanted into the host cortex migrated toward the host cortical surface, sometimes in several radial lines. Astrocytes from grafts, especially callosal grafts, placed in the subcortical white matter migrated along the host fiber tracts. Many astrocytes transplanted into the hippocampus formed laminar patterns close to the hippocampal neuronal layers. These results suggest that the direction, pattern, and speed of astrocyte migration are influenced by local substrates in the host brain.

Nucleolar changes in senescing WI-38 cells.

Changes in the area, dry mass and morphology of nucleoli were studied during in vitro aging of WI-38 cells. Interferometric methods were used for nucleolar dry mass determinations. The results show that there is (1) an increase in the fraction of cells with one large nucleolus per nucleus, 17% at population doubling 27.3 vs. 93% at population doubling 41.2, (2) an increase in mean nucleolar dry mass (583% at the last doubling), and (3) an increase in mean nucleolar area (236% at the last doubling) with in vitro senescence of WI-38 fibroblast cells. A strong correlation (r = 0.92) between nucleolar dry mass and nucleolar area was demonstrated.

Characterization of avian reovirus non structural protein sigmaNS synthesized in Escherichia coli.

The coding region of avian reovirus S1133 genomic segment S4, encoding the non structural protein sigmaNS, was inserted into expression vector pET28a and the protein was expressed in Escherichia coli BL21(DE3) as a fusion protein containing a C-terminal peptide with six tandem histidines (His-tag). The expressed protein (esigmaNS) consistent with the expected molecular size of the avian reovirus protein sigmaNS synthesized in infected cells was readily purified by His-Bind Resin. The esigmaNS was further confirmed to be indistinguishable from viral sigmaNS by immunoblot analysis. The esigmaNS binds 32P-labeled ssRNA probe produced by run-off transcription of clone pGEM-3Zf(+)S4. The binding activity is blocked by heterologous yeast rRNA, but not by homologous avian reovirus dsRNA and heterologous infectious bursal disease virus dsRNA and salmon sperm dsDNA. Therefore, the ssRNA-binding activity of the expressed protein sigmaNS is non sequence-specific, similar to that previously described for viral sigmaNS purified from avian reovirus infected cell extracts. In addition, the recent data also show that the optimal salt (NaCl) concentration and pH for its binding are 100-150 mM and 7.0, respectively, in terms of the UV cross-linking and RNase A treatment of the reaction mixtures prior to the denaturing gel analysis.

The role of aspiration biopsy cytology in the diagnosis of pulmonary tuberculosis.

The experience at one institution concerning the diagnosis of pulmonary tuberculosis by aspiration biopsy is reviewed. Twelve cases in which acid-fast bacilli were identified in pulmonary aspiration material are reported. The cytologic findings were confirmed by microbiologic culture in nine cases. The advantages of using aspiration biopsy cytology in the diagnosis of pulmonary tuberculosis are discussed. In our experience we conclude that the use of aspiration biopsy cytology in the diagnosis of pulmonary tuberculosis is not only accurate, but also offers several advantages that are important in patient care.

Markers of thrombin and plasmin generation in patients with inherited thrombophilia.

AIM: To determine the prevalence of a biochemically detectable hypercoagulable state, defined in terms of increased thrombin or plasmin generation, in patients with phenotypically characterised thrombophilia. METHODS: Plasma concentrations of the prothrombin activation peptide F1.2 and fibrin degradation (FbDP) and fibrinogen degradation products (FgDP) were measured by enzyme immunoassay in 104 patients deficient in natural anticoagulants, and 35 unaffected relatives. RESULTS: Increased concentrations of F1.2, FbDP, and FgDP were present in 18, 25, and 19 of 104 patients, respectively. There were no correlations between F1.2, FbDP, and FgDP concentrations, or between these parameters and concentrations of natural anticoagulants except for a negative correlation between protein C concentrations and FgDP (rho = -0.46, p = 0.009). CONCLUSION: A biochemically detectable hypercoagulable state is present in some patients with asymptomatic thrombophilia. Markers of plasmin generation may be increased more frequently in thrombophilia than markers of thrombin generation. This finding should prompt the inclusion of markers of plasmin generation in prospective longitudinal cohort studies to determine the predictive value of a hypercoagulable state, defined by either excessive thrombin or plasmin generation, for the development of venous thromboembolism.

Characterization of nonradioactive hybridization probes for detecting infectious bursal disease virus.

Reverse transcription followed by the polymerase chain reaction was used to amplify a fragment of infectious bursal disease virus (IBDV) strain P3009 genome. The amplified DNA fragment was annealed into the plasmid pUC18 and used to transform Escherichia coli strain JM109. A clone that contained IBDV-specific nucleotide sequences was selected and designated pC23. The DNA fragment within pC23 was 320 base pairs in length and designated C23. Radiolabeled probes prepared from C23 hybridized to genome segment A of strain P3009 by a northern-blot hybridization assay. Biotin-labeled probes prepared from C23 and pC23 either by using nick translation (designated C23/NT and pC23/NT, respectively) or by direct introduction of biotin molecules into C23 and pC32 (designated C23/BH and pC23/BH, respectively) were used in the dot blot hybridization assay for detecting IBDV strains. All four biotinylated probes detected three serotype 1 viruses and one serotype 2 IBDV. However, they did not cross-react with nucleic acids extracted from mock-infected cells or from seven unrelated avian viruses. Probe pC23/BH detected as little as 0.04 ng of IBDV RNA, while the other three probes were less sensitive and detected approximately 1 ng of IBDV RNA. In addition, the probe pC23/BH detected IBDV RNA in bursa tissues from commercial broiler raising farms following the dot blot hybridization.

A monoclonal antibody capture enzyme-linked immunosorbent assay for detecting antibodies to infectious bursal disease virus.

A monoclonal antibody capture enzyme-linked immunosorbent assay (mAb-ELISA) for antibodies to infectious bursal disease virus (IBDV) in chicken sera was developed and compared with conventional ELISA. When sera from farm chickens were tested by the two ELISAs and serum neutralization (SN), the correlation rate between SN and mAb-ELISA was 100% (49/49), and that between SN and conventional ELISA was 81.6% (40/49). In mAb-ELISA, all of the sera that were antibody-negative by SN had low absorbance values (below 0.05), and the absorbance values correlated closely with the SN titers. In the conventional ELISA, however, the sera antibody-negative by SN had various absorbance values ranging from 0.06 to 0.32. mAb-ELISA had much lower non-specific reactions than the conventional ELISA against sera from IBD-negative chickens.