High-grade B-cell lymphoma, not otherwise specified: a multi-institutional retrospective study.

In this multi-institutional retrospective study, we examined the characteristics and outcomes of 160 patients with high-grade B-cell lymphoma, not otherwise specified (HGBL-NOS)-a rare category defined by high-grade morphologic features and lack of MYC rearrangements with BCL2 and/or BCL6 rearrangements ("double hit"). Our results show that HGBL-NOS tumors are heterogeneous: 83% of patients had a germinal center B-cell immunophenotype, 37% a dual-expressor immunophenotype (MYC and BCL2 expression), 28% MYC rearrangement, 13% BCL2 rearrangement, and 11% BCL6 rearrangement. Most patients presented with stage IV disease, a high serum lactate dehydrogenase, and other high-risk clinical factors. Most frequent first-line regimens included dose-adjusted cyclophosphamide, doxorubicin, vincristine, and etoposide, with rituximab and prednisone (DA-EPOCH-R; 43%); rituximab, cyclophosphamide, doxorubicin, vincristine, and prednisone (R-CHOP; 33%); or other intensive chemotherapy programs. We found no significant differences in the rates of complete response (CR), progression-free survival (PFS), or overall survival (OS) between these chemotherapy regimens. CR was attained by 69% of patients. PFS at 2 years was 55.2% and OS was 68.1%. In a multivariable model, the main prognostic factors for PFS and OS were poor performance status, lactate dehydrogenase >3 × upper limit of normal, and a dual-expressor immunophenotype. Age >60 years or presence of MYC rearrangement were not prognostic, but patients with TP53 alterations had a dismal PFS. Presence of MYC rearrangement was not predictive of better PFS in patients treated with DA-EPOCH-R vs R-CHOP. Improvements in the diagnostic criteria and therapeutic approaches beyond dose-intense chemotherapy are needed to overcome the unfavorable prognosis of patients with HGBL-NOS.

Molecular detection of Chlamydophila abortus, Coxiella burnetii, and Mycoplasma agalactiae in small ruminants' aborted fetuses in southern Iran.

Abortion in sheep and goats has become increasingly important worldwide because of the significant economic losses and potential zoonotic implication of commonly involved pathogens. Therefore, this cross-sectional study was conducted in southern Iran to detect the Chlamydophila abortus and Coxiella burnetii, as zoonotic pathogens, and Mycoplasma agalactiae, as a neglected abortifacient agent in small ruminants' aborted fetuses, by using polymerase chain reaction (PCR). From a total of 300 aborted fetuses (183 sheep and 117 goats), 46 samples (15.5%) were positive by PCR, 11% for C. abortus, 2% for C. burnetii, and 3% for M. agalactiae. Also, the association of suggested risk factors with abortion due to these bacterial agents was investigated using univariable and multivariable logistic regression. Results of the statistical analysis showed significant association of C. abortus with flock size (OR = 2.82, P = 0.014), season (P < 0.05), and the number of pregnancy in the aborted dam (OR = 2.5, P = 0.05). Our results indicated that C. abortus has a relatively substantial role in small ruminant abortions, and C. burnetii and M. agalactiae are likely important abortifacient agents in our region, too. Regarding veterinary and/or public health importance of these bacterial agents, more attention from veterinary and/or human health services and, maybe, a surveillance system for control and prevention of them are recommended.

Histologic and histomorphometric changes of testis following oral exposure to methyl tertiary-butyl ether in adult rat.

Methyl tertiary-butyl ether (MTBE) is used to reduce carbon monoxide and ozone in urban air and to boost fuel octane. There is a lack of knowledge in the literature about the histomorphometric changes of the testis following exposure to MTBE. Therefore, this experimental study was performed to determine the effect of oral exposure to MTBE on histologic and histomorphometric changes of testis in adult rat. A total of 25 adult male Sprague-Dawley rats were randomly divided into five equal experimental groups: control, almond oil and three treatment groups which received 400, 800 and 1600 mg/kg/day MTBE in almond oil by gavages for 30 consecutive days. Histomorphometric analysis showed no significant difference in absolute and relative testis weight, connective tissue thickness, germinal epithelium height, tunica albuginea thickness and Sertoli cell numbers between experimental groups (P>0.05). However, trend analysis showed that the seminiferous tubule diameter increased and interstitial cell numbers as well as spermatocyte and spermatid cell numbers decreased significantly in MTBE treated groups (P<0.05). It may be concluded that MTBE could exert adverse effects on spermatogenic cells in adult rat. Whether the observed changes in the present study are due to the direct effect of MTBE via passing blood-testis barrier or its indirect effect through another mechanism should be elucidated in future studies.

Relationships between thyroid hormones and serum energy metabolites with different patterns of postpartum luteal activity in high-producing dairy cows.

This study investigated the relationships of thyroid hormones, serum energy metabolites, reproductive parameters, milk yield and body condition score with the different patterns of postpartum luteal activity in the postpartum period. A total of 75 multiparous healthy (free of detectable reproductive disorders) Holstein dairy cows (mean peak milk yield = 56.5 ± 7.0 kg/day) were used in this study. Transrectal ultrasound scanning and blood sample collection were performed twice weekly. Serum concentrations of progesterone (P4) were measured twice weekly and beta-hydroxybutyrate (BHBA), non-esterified fatty acids, thyroxine (T4), 3,30,5-tri-iodothyronine (T3), free thyroxine (fT4) and free 3,30,5-tri-iodothyronine (fT3) were measured every 2 weeks from the 1st to the 8th week postpartum. On the basis of the serum P4 profile of the cows, 25 (33.4%) had normal luteal activity (NLA), whereas 30 (40%), 10 (13.3%), 6 (8%) and 4 (5.3%) had prolonged luteal phase (PLP), delayed first ovulation (DOV), anovulation (AOV) and short luteal phase, respectively. Serum T4 concentrations in PLP cows were higher than that in NLA cows at the 3rd week postpartum and did not change during the period of study, whereas in the NLA cows the concentrations increased (P < 0.05). Further, the least square (LS) mean of serum fT4 concentrations in the DOV and AOV cows were significantly lower than in the NLA cows during the study period (P < 0.05). In addition, the AOV cows had higher LS mean serum BHBA and T4 concentrations than the NLA cows in early weeks postpartum (P < 0.05). In conclusion, the serum thyroid hormones' profile differs in high-producing dairy cows showing PLP, AOV and DOV in comparison with the postpartum NLA cows.

Evaluation of sialic acid and acute-phase proteins (haptoglobin and serum amyloids A) in healthy and avian infection bronchitis virus-infected chicks.

Forty-five 24-day-old Cobb chicks infected with infectious bronchitis virus (IBV) and ten healthy 24-day-old Cobb chicks without any clinical signs of IBV as control group were selected for the study. All of the diseased chicks showed some or all of the clinical signs of infectious bronchitis including gasping, coughing and nasal discharge, wet eyes, swollen sinuses, reduction of food consumption and weight gain. Diagnosis of IBV was based on clinical signs and ELISA test. Blood samples were taken from the wing vein into two tubes: one containing ethylenediaminetetraacetic acid (EDTA) and one without EDTA. Haptoglobin (Hp), serum amyloid A (SAA), total sialic acid (TSA), lipid-bound sialic acid (LBSA) and protein-bound sialic acid (PBSA) concentrations were measured. All of the study variables were significantly higher in diseased birds compared with control group. Results showed that there were significant positive correlations between TSA, LBSA and PBSA in both groups. No correlation was observed between Hp and SAA with any other parameters; however, there was significant negative correlation between Hp and SAA in the control group. Results for receiver operating characteristic analysis showed that area under the curve (AUC) for TSA, LBSA, PBSA, Hp and SAA were 0.93, 0.98, 0.90, 0.90 and 0.80, respectively. According to AUC, LBSA was the most sensitive factor to change in the diseased birds. It can be concluded that in naturally occurring IBV infection, significant increases in TSA, LBSA, PBSA, Hp and SAA concentrations are expected and among study variables, LBSA had the most obvious change so it may be considered as the most sensitive parameter.

DRAQ5-based, no-lyse, no-wash bone marrow aspirate evaluation by flow cytometry.

Flow cytometry (FC) is a powerful tool for objective phenotyping of hematolymphoid neoplasia. Analysis of bone marrow aspirates and peripheral blood specimens by FC typically requires an erythrocyte lysis or gradient separation method to remove erythrocytes prior to analysis, which may result in the loss of certain populations, in particular nucleated erythroid cells. We developed a method to analyze bone marrow aspirates (BMAs) by FC without erythrocyte lysis or washing to minimize cell loss by exploiting the nuclear DRAQ5 fluorescence as a gating parameter (DRAQ5 protocol). We analyzed a total of 31 BMAs from patients with a variety of diagnoses utilizing the DRAQ5 protocol in combination with CD71 and CD45 antibodies to determine the marrow differentials. These were compared with differential counts obtained by morphologic study and erythrocyte lysis FC. The DRAQ5 protocol preserved the nucleated erythrocytes, allowing calculations of the myeloid to erythroid ratio and of blasts/abnormal cells that better reflect the morphologic nucleated cell differential than erythrocyte lysis FC.

Acute bilineal leukemia: a rare disease with poor outcome.

Most cases of acute leukemia can be assigned to the myeloid, B or T lineage. In a few cases, definitive assignment cannot be achieved because blasts express antigens of more than one lineage. A subset of these, referred to as acute bilineal leukemias (aBLLs), is characterized by the presence of more than one population of blasts, each comprising a single lineage. We identified 19 cases of aBLL, including 10 mixed T and myeloid (T-My) and nine mixed B and myeloid (B-My); no mixed B and T cases were identified. Cytogenetic data were available for 16 patients. Three of seven patients with B-My had a t(9;22)(q34q11.2), two had 11q23 translocations and one had del(9). Two of nine patients with T-My had 2p13 translocations; five had other unrelated abnormalities. Of 16 patients with outcome data, only six achieved complete remission and only two remain free of disease 2.5 and 4.5 years after chemotherapy or stem cell transplantation. aBLL is a rare disease that combines B or T and myeloid blasts. Cytogenetic abnormalities of t(9;22) and 11q23 are common in, and may be restricted to, B-My cases, while T-My cases have frequent but generally non-recurring abnormalities. Both types of aBLL are associated with poor outcome.

Antibody response of chickens to serotype 1, 2, or 3 Marek's disease vaccines based on ELISA with infected cells as antigen.

An enzyme-linked immunosorbent assay (ELISA) was applied to evaluate the antibody response of commercial White Leghorn chickens to vaccination against Marek's disease (MD) at hatch (day 0) with serotype-1 (Rispens), -2 (SB-1), or -3 (turkey herpesvirus, HVT) vaccine virus and to challenge on day 21 with MD virus. Antigens for the test were whole chicken embryo fibroblast cells infected with Rispens, SB-1, or HVT. The chickens were progeny of stock that had been vaccinated with HVT, and on day 21 the nonvaccinated group had higher levels of maternal antibodies to HVT than to other antigens (P < 0.05). Only SB-1 vaccine had induced antibodies by day 21, and this was detected only against homologous antigens. On day 49, all three vaccines had induced higher levels of antibodies to homologous than to heterologous antigens. Marek's Disease virus (MDV) induced antibodies to all three antigens, but challenging vaccinated chicks did not significantly increase levels of antibodies on day 81 to any of the three antigens. It was concluded that an ELISA using whole cells as antigens would have potential value for monitoring the antibody response induced by MD vaccines and virulent MDV.

The DNA sequence, annotation and analysis of human chromosome 3.

After the completion of a draft human genome sequence, the International Human Genome Sequencing Consortium has proceeded to finish and annotate each of the 24 chromosomes comprising the human genome. Here we describe the sequencing and analysis of human chromosome 3, one of the largest human chromosomes. Chromosome 3 comprises just four contigs, one of which currently represents the longest unbroken stretch of finished DNA sequence known so far. The chromosome is remarkable in having the lowest rate of segmental duplication in the genome. It also includes a chemokine receptor gene cluster as well as numerous loci involved in multiple human cancers such as the gene encoding FHIT, which contains the most common constitutive fragile site in the genome, FRA3B. Using genomic sequence from chimpanzee and rhesus macaque, we were able to characterize the breakpoints defining a large pericentric inversion that occurred some time after the split of Homininae from Ponginae, and propose an evolutionary history of the inversion.

The finished DNA sequence of human chromosome 12.

Human chromosome 12 contains more than 1,400 coding genes and 487 loci that have been directly implicated in human disease. The q arm of chromosome 12 contains one of the largest blocks of linkage disequilibrium found in the human genome. Here we present the finished sequence of human chromosome 12, which has been finished to high quality and spans approximately 132 megabases, representing approximately 4.5% of the human genome. Alignment of the human chromosome 12 sequence across vertebrates reveals the origin of individual segments in chicken, and a unique history of rearrangement through rodent and primate lineages. The rate of base substitutions in recent evolutionary history shows an overall slowing in hominids compared with primates and rodents.

The DNA sequence of the human X chromosome.

The human X chromosome has a unique biology that was shaped by its evolution as the sex chromosome shared by males and females. We have determined 99.3% of the euchromatic sequence of the X chromosome. Our analysis illustrates the autosomal origin of the mammalian sex chromosomes, the stepwise process that led to the progressive loss of recombination between X and Y, and the extent of subsequent degradation of the Y chromosome. LINE1 repeat elements cover one-third of the X chromosome, with a distribution that is consistent with their proposed role as way stations in the process of X-chromosome inactivation. We found 1,098 genes in the sequence, of which 99 encode proteins expressed in testis and in various tumour types. A disproportionately high number of mendelian diseases are documented for the X chromosome. Of this number, 168 have been explained by mutations in 113 X-linked genes, which in many cases were characterized with the aid of the DNA sequence.

FLT3 mutations in myeloid sarcoma.

Myeloid sarcoma is an extramedullary tumour that typically occurs in the setting of acute myeloid leukaemia (AML), or myeloproliferative disorders. In AML, two types of mutations in Fms-like tyrosine kinase 3 (FLT3) have been described; internal tandem duplications (ITD) and point mutations at aspartic acid residue 835 (D835). We analysed 24 myeloid sarcoma specimens from 20 patients for FLT3 ITD and D835 mutations. FLT3 ITD mutations were identified in three of 20 cases (15%); no D835 mutations were identified. The ITD inserts ranged in size from 33 to 198 base pairs (bp) and represented approximately 20-40% of the FLT3 alleles. Two cases showed discordance in FLT3 ITD mutational status. In one case, the leukaemia specimen was positive for a FLT3 ITD mutation and the myeloid sarcoma specimen was negative. In the second case, the myeloid sarcoma was positive for a FLT3 ITD mutation at diagnosis, but negative in subsequent relapse samples. Our findings suggest that small molecule inhibitors of FLT3 may be useful therapeutic agents for treatment of myeloid sarcomas-containing FLT3 mutations, however, the potential for discordance between the leukaemia and myeloid sarcoma, necessitates that the myeloid sarcoma tumour itself be analysed for FLT3 mutations.

Consanguineous marriage in Iran.

Consanguineous marriage is a major feature of family systems in south-west Asia. The aim of the present study was to determine the current prevalence and patterns of consanguinity in Iran as a means of assessing the associated requirement for genetic counselling services. Consanguinity was studied in 12 ethnic/religious populations, the Persians (Shi'a and Sunni), Kurds (Shi'a and Sunni), Lurs, Azaris, Baluchis, Zabolis, Turkamans, Bakhtiaris, Ghashghais and Arabs. A multi-stage sampling design was used with a representative total sample of 306 343 couples. The overall rate of consanguineous marriage was 38.6% with a mean inbreeding coefficient (alpha) of 0.0185. First cousin marriages (27.9%) were the most common form of consanguineous union, with parallel patrilateral marriage especially favoured. Statistically significant differences were observed in the prevalence and patterns of consanguinity between ethnic/religious populations and geographical regions. There also were significant differences for proportions of consanguineous marriages between Shi'a and Sunni populations within the same ethnic group. The highest rates of consanguineous union were in the least affluent sections of the population.

Influence of GSTT1 null genotype on the offspring sex ratio of gasoline filling station workers.

The aim of this study was to investigate whether the occupational exposure to gasoline of men employed at filling stations affects the sex of their children. Altogether 115 offspring (47 males, 68 females) were identified within families of 49 men working in filling stations in Shiraz (Fars province, south of Iran) and 345 offspring (178 males, 167 females) from 147 families of unexposed persons from the general population of Shiraz, which were matched by age of fathers (+/-2 years) and number of children as a control group. The offspring sex ratio at birth (male proportion) in the filling station workers was significantly lower than the ratio in control group (OR = 0.65; 95% confidence intervals (CI) 0.42 to 0.99). Genotypes of glutathione S-transferase M1 (GSTM1) and T1 (GSTT1) were investigated on extracted genomic DNA of 37 exposed workers using the polymerase chain reaction based method. In exposed group with active GSTM1 and GSTT1 genes, offspring sex ratio was the same as the ratio in the control group (OR = 0.66; 95% CI 0.34 to 1.28). However, in the exposed group with active GSTM1 and null genotype of GSTT1, the offspring sex ratio statistically decreased (OR = 0.45; 95% CI = 0.21 to 0.96). It seems that the GSTT1 null genotype has an effect on offspring sex ratio in the filling station workers.

Distinction of endocervical and endometrial adenocarcinomas: immunohistochemical p16 expression correlated with human papillomavirus (HPV) DNA detection.

Determining the origin of uterine adenocarcinomas can be difficult in biopsy and curettage specimens because the morphologic spectrum of endocervical and endometrial adenocarcinomas overlaps. In hysterectomy specimens, the primary site is often equivocal for tumors that involve the lower uterine segment and endocervix and lack identifiable precursor lesions. Most endocervical adenocarcinomas (ECAs) contain high-risk human papillomavirus (HPV) DNA, whereas endometrial adenocarcinomas (EMAs) rarely do. p16 is an inhibitor ofcyclin-dependent kinases, and overexpression of p16 has been observed in cervical intraepithelial lesions and invasive carcinomas associated with high-risk HPV types. We evaluated the utility of immunohistochemistry for p16 in the distinction of ECAs and EMAs. p16 expression was assessed in 24 unequivocal EMAs and 19 unequivocal ECAs and correlated with HPV DNA detection by in situ hybridization and polymerase chain reaction. These assays were then used to assist in the classification of four lower uterine segment/upper endocervical adenocarcinomas (LUS/EC-A) of equivocal origin. p16 expression was moderate-strong and diffuse in 18 ECAs (median 90% of tumor cells positive, range 90%-100%), and weak and diffuse in one. Fourteen of these were positive for HPV DNA, whereas 5 lacked detectable HPV DNA by in situ hybridization; one of these 5 was positive by polymerase chain reaction. In contrast, EMAs displayed weaker staining with patchy distribution (median 30% of tumor cells positive, range 5%-70%) and none contained HPV DNA by in situ hybridization. Two LUS/EC-As, which were positive for HPV, exhibited strong, diffuse p16 expression, consistent with endocervical origin of the tumors. The remaining 2 LUS/EC-As showed patchy p16 staining and did not contain detectable HPV DNA, consistent with the endometrial origin of the tumors. The p16 expression pattern can distinguish ECAs from EMAs. Compared with HPV DNA detection by in situ hybridization, p16 immunohistochemistry appears to be a more sensitive and easier to perform method for distinguishing ECAs from EMAs, can be used to assist in the classification of LUS/EC-As of equivocal origin, and should be evaluated for its utility in the prospective classification of uterine adenocarcinomas in curettage specimens prior to hysterectomy.

Fine-needle aspiration of abdominal fat pad for amyloid detection: a clinically useful test?

Fine-needle aspiration of abdominal fat pad (FNAFP) is commonly employed for the diagnosis of systemic amyloidosis, a disease with highly variable clinical manifestations, often presenting difficult patient management problems. We evaluated the role of FNAFP particularly in reference to its clinical usefulness. Pathology reports and clinical histories of 91 consecutive cases of FNAFP with Congo red (CR) staining at The Johns Hopkins Hospital (1999-2000) were reviewed. Major emphases were assessment of the clinical utility of the test, correlation with concurrent or subsequent biopsies, and treatment strategies. The primary indications for FNAFP were monoclonal gammopathy (34%), cardiomyopathy (22%), renal insufficiency (20%), neuropathy (8%), plasma cell dyscrasia (6%), and other conditions (10%). Of the 91 patients who underwent FNAFP, the results were as follows; 20 cases (22%) positive; 62 cases (68%) negative; eight cases (9%) insufficient for diagnosis; and one case (1%) equivocal. Of the 20 positive cases, follow-up biopsies were performed on 11 cases, of which six were positive and five were negative for amyloid by CR. Of the 62 negative cases, follow-up biopsies were performed on 19 cases, 14 of which were negative and five positive for amyloid by CR. A follow-up biopsy on the single equivocal case was positive for amyloid by CR. Twenty-one patients positive for amyloid, based on initial or follow-up biopsies, were managed symptomatically without any specific treatment for amyloidosis. One patient, who was specifically treated for amyloidosis by melphalan and dexamethasone, died 1 wk after therapy. Three patients with multiple myeloma and amyloidosis underwent chemotherapy. We conclude that primary clinical indications for FNAFP for amyloidosis are highly variable. An FNAFP result is often not considered clinically conclusive and is followed by further invasive procedures to detect amyloid (55% of our positive and 31% of our negative FNAFP cases were rebiopsied). The estimated sensitivity and specificity of FNAFP were 75% and 92%, respectively. Overall, the reliance on the results of FNAFP depended on the degree of clinical suspicion of the treating physician. Although in the majority of cases diagnosis of amyloidosis did not alter the treatment strategies, a conclusive positive result helped in ruling out other underlying conditions as the cause of patients' symptoms.

Detection of human papillomavirus in cervical lymph nodes: a highly effective strategy for localizing site of tumor origin.

PURPOSE: Patients with head and neck squamous cell carcinoma (HNSCC) often come to clinical attention with a neck mass due to metastatic spread to lymph nodes. Treatment is dictated by the subsequent determination of primary tumor site and stage. However, the primary site remains elusive in some patients even after an exhaustive examination. Human papillomavirus type 16 (HPV-16) is an important etiologic agent for HNSCCs that arise within the oropharynx but less so for tumors from nonoropharyngeal sites. Detection of HPV-16 or a surrogate marker may be useful in localizing tumor origin in patients who present with metastatic HNSCC. EXPERIMENTAL DESIGN: We performed in situ hybridization (ISH) for HPV-16 on lymph node metastases from 68 patients with HNSCC. P16 immunohistochemistry was also performed because HPV-16 integration disrupts the retinoblastoma pathway and induces an overexpression of p16. RESULTS: HPV-16 was detected in 22 of the 68 (32%) cases by ISH. When stratified by site of origin, HPV-16 was detected in 22 of 31 (71%) metastases from the oropharynx, but in none of the 37 (0%) metastases from other sites (P < 0.001; Fisher's exact). P16 expression was associated with the presence of HPV-16 by ISH: 21 of 22 HPV-16 positive tumors exhibited p16 expression, whereas only 4 of the 46 HPV-16-negative tumors were p16 positive (95% versus 9%; P < 0.001; Fisher's exact). P16 expression in the node metastases also correlated with site of tumor origin: 24 of 31 oropharyngeal tumors were p16 positive, whereas only 1 of 37 nonoropharyngeal tumors was p16 positive (77% versus 3%; P < 0.001; Fisher's exact). CONCLUSIONS: For patients with metastatic HNSCC, detection of HPV-16 is a reliable way to establish origin from the oropharynx, either directly by ISH or indirectly by immunohistochemistry for p16 overexpression.

Immature granulocyte measurement using the Sysmex XE-2100. Relationship to infection and sepsis.

We determined the usefulness of immature granulocyte measurement as a predictor of infection or positive blood culture and compared the results with total WBC count and absolute neutrophil count (ANC). Blood samples from 102 infected and 69 noninfected patients were analyzed using the Sysmex XE-2100 automated blood cell counter (Sysmex, Kobe, Japan). The percentage of immature granulocytes was significantly higher in infected than in noninfected patients and in patients with positive than patients with negative blood cultures. Receiver operating characteristic curves showed that the percentage of immature granulocytes was a better predictor of infection than the WBC count and comparable to the ANC. Automated immature granulocyte measurements reflect a biologically and clinically relevant phenomenon but are not sensitive enough to be used as screening assays for prediction of infection or bacteremia. However, although infrequently encountered, a percentage of immature granulocytes of more than 3 was a very specific predictor of sepsis and might help expedite microbiologic laboratory evaluation of a subset of patients.