Post-mortem computed tomography differentiation of putrefactive gas and air embolism: A case of traumatic carotid-jugular arteriovenous fistula.

INTRODUCTION: PMCT is superior to autopsy for identification of intravascular or extravascular gas pockets and their distribution. However, differentiation between air embolism and putrefactive gas can prove challenging due to overlapping imaging findings. CASE REPORT: We report a case of a healthy young man who was involved in a fight, sustained a slash wound to the right side of his head by a kitchen knife and died at the scene. Pre-autopsy PMCT demonstrated complex fractures of the right mastoid bone extending to the right petrous apex and jugular bulb, exposing the right sigmoid sinus. There was also asymmetric intravascular air distribution suspicious of air embolism with ancillary findings of traumatic carotid-jugular pseudoaneurysm and arteriovenous fistulous formation. Post-mortem examination revealed a slash wound measuring 12x2 cm at the right side of the head, cutting through the scalp, right temporal bone, right temporal meninges, right sigmoid venous sinus and part of the right occipital lobe. No intracranial haemorrhage was found on both PMCT and autopsy. DISCUSSION: PMCT findings of air embolism versus putrefactive air on PMCT are discussed in this case. Detailed history on mechanism, circumstances, time of death and careful analysis of intravascular and extravascular air distribution patterns on PMCT are essential in guiding differentiation of true fatal air embolism and "normal" post-mortem putrefactive air. Needless to say, it is recommended that PMCT be performed as early as possible after death to reduce the chances and presence of artifactual decomposition changes.

Establishment of effective and safe recipient preparation for germ-cell transplantation with intra-testicular busulfan treatment in pre-pubertal Barbari goats.

The busulfan, an alkylating agent, suppresses endogenous spermatogenesis in recipient testes. However, considering a wide variation in the effects of busulfan among animal species, its dosage and route of infusion need optimization to prepare effective and safe recipients. Thus, the current study aimed to create a suitable recipient goat model for germ cell (Gc) transplantation through a single intra-testicular (i.t.) busulfan infusion under ultrasonographic (USG) guidance. As observed through the infusion of trypan blue under USG guidance into mediastinum testis (MT) of pre-pubertal Barbari bucks, 3-5 mL of trypan blue solution could fill almost 80% of seminiferous tubules. Thereafter, in Experiment-1, the effect of different busulfan doses (mg/kg) i.e. 0 [negative control, Group (Gr) 1; 0 mg/kg-MT], 1 (Gr 2; 1 mg/kg-MT), 2 (Gr 3; 2 mg/kg-MT), and 3 (Gr 4; 3 mg/kg-MT) were studied. Further, in Experiment-2, sterilizing effects of busulfan infusion through two different routes [MT or cavum vaginale (CV)] were compared. Following i.t. busulfan treatment, no adverse physiological effects or body weight loss were detected. The histological analyses demonstrate a dose-dependent depletion of Gc with almost complete loss of Gc and spermatogenic activities in Gr 3 and 4, and extensive fibrosis in Gr 4. A considerable suppression of spermatogenesis marked with devoid of endogenous spermatogonial population and absence of significant (P > 0.05) effect on key hematological variables were observed in 2 mg/kg-MT Gr. These findings coupled with the results of significant (P < 0.05) down-regulation of marker genes of undifferentiated spermatogonia (THY-1 and PLZF), Gc pluripotency (UCHL-1, OCT-4, and DDX-4), and adhesion (E-cadherin and ß-integrin); up-regulation of apoptotic genes (ID - 4 and BCL-6), and unchanged expression of Sertoli cell marker (vimentin), confirmed the safe and efficient depletion of endogenous Gc in 2 mg/kg-MT Gr. Furthermore, the effect of busulfan infusion on scrotal-testicular biometry, endocrine variables (plasma cortisol and testosterone), and Gc removal was more evident when busulfan was infused into MT than into CV. Overall, the results demonstrated that 2.0 mg/kg is an optimal single dose of busulfan when infused into the MT under USG guidance for the preparation of pre-pubertal recipient bucks. Overall, this study provides a basis to prepare suitable recipients through providing an available niche for efficient Gc transplantation in goats.

Immunophenotypic Characterization and Ploidy Analysis of Neoplastic Plasma Cells by Multiparametric Flow Cytometry.

Flow cytometric (FCM) immunophenotyping is an important tool for generating diagnostic and prognostic information in plasma cell dyscrasias. This study aimed to evaluate the immunophenotype and ploidy status of plasma cells (PCs) in patients of myeloma and its correlation with other laboratory parameters. Bone marrow of 70 newly diagnosed cases of myeloma were subjected to FCM using a panel of antibodies; CD138, CD38, CD19, CD45, CD28, CD81, CD56, CD200, and CD229. FxCycle Violet (FCV) dye was used for the ploidy analysis of clonal PCs. Median age was 60 years with M:F ratio of 3.2:1. A positive correlation was noted between the morphological and FCM-based PC enumeration (r = 0.4, p = 0.001). Aberrant expression of CD56, CD200, CD28, CD117, CD81 and CD19 and was observed in 88.5%, 77%, 29%, 37%, 23% and 17% cases respectively. Two aberrant antigens were noted in all cases. CD81 + cases had a relatively higher quantity of monoclonal-protein (> 1 g/dl, p < 0.05) and renal insufficiency (Cr > 2 mg/dl, p < 0.05) as compared to the CD81- cases. CD229 was expressed in all the cases, with a median MFI in PCs significantly higher than other hematopoietic elements. Hyperdiploid PCs (median DI-1.59, range, 1.16-2.6) were noted in 80% cases (n = 48), diploid/ near-hyperdiploid PCs in 8% (n = 5) cases and hypodiploidy in 3% (n = 1) cases. Bright CD56/CD200 and CD45- can identify abnormal PC in the majority of the cases. CD81 appears to correlate with disease burden and might be useful as a prognostic marker. CD229 is a reliable gating marker for plasma cells. Ploidy analysis may be incorporated in routine workup to guide in the identification of patients with poor prognosis. Supplementary Information: The online version contains supplementary material available at 10.1007/s12288-021-01477-y.

Prevalence of incidental and total human immunodeficiency virus, hepatitis B and hepatitis C seropositivity among patients posted for cataract surgery at a tertiary care center in India.

PURPOSE: To study the prevalence of human immunodeficiency virus (HIV), hepatitis B (HBV), and hepatitis C (HCV) viral seropositive among the patients posted for cataract surgery at a tertiary care center in north India. METHODS: It was a cross-sectional study done for 30 months duration. All the patients posted for cataract surgery underwent comprehensive ophthalmic evaluation followed by routine hematological workup, including viral markers for HIV, Hepatitis B surface antigen (HBsAg), and anti-HCV. Data were analyzed by the Statistical Package for Social Science (SPSS Version 20). RESULTS: A total of 7,316 individuals underwent cataract surgery from Jan 2016 to August 2018, 4,073/7,316 (55.7%) were males. The prevalence for HIV was 58/7,316 (0.8%), HBsAg was 151/7,316 (2.1%), and HCV was 11/7,316 (0.1%); 28/58 (48.3%) HIV positives were unaware of their seropositivity till testing, as were 37/151 (24.5%) of HBsAg positives, and 4/11 (36.4%) HCV positives. There was a significant relationship between the mean age in the patients with HIV (P = 0.002) and anti-HCV (P = 0.045). A majority of the seropositive patients were found to be illiterate (45.6%), followed by educated up to high school level (29.1%), and graduate (25.0%). CONCLUSION: Viral seropositivity was significant among the patients posted for cataract surgery. The eye care providers could refer these patients for counseling and further management for the patient's and their caretaker's benefit.

Association of TYRP1 with hypoxia and its correlation with patient outcome in uveal melanoma.

PURPOSE: Molecular mechanisms of uveal melanoma development in association with high pigmentation are unclear. Tyrosinase Related Protein (TYRP1) is not only one of the important melanogenesis marker that contributes to melanin synthesis, but can also prevents the melanocyte death. The induction of melanogenesis leads to induction of HIF-1α which can affect the behavior of melanoma cells and its surrounding environment. The aim of our study was to determine the expression of TYRP1 and HIF-1α at the protein and RNA level and determine its prognostic significance. METHODS: In the present study, the expression of TYRP1 and HIF-1α was investigated on 61 formalin-fixed paraffin-embedded choroidal melanoma samples by immunohistochemistry. Fresh 50 samples were validated by real-time PCR. Results were correlated with clinicopathological parameters and Kaplan-Meier was performed to determine the prognostic significance. RESULTS: High immunoexpression of TYRP1 and HIF-1α was present in 61 and 54% of patients, respectively. Both TYRP1 and HIF-1α correlated well with high pigmentation and BAP1 (BRCA1 Associated Protein-1) loss (p < 0.05) at IHC level as well as transcriptional level. There was reduced metastatic free survival in patients with necrosis and this was statistically significant (p = 0.010). CONCLUSION: Our findings indicate that TYRP1 can be used as a potential biomarker in the development of targeted therapy in UM. Further studies on melanogenesis markers associated with TYRP1 could provide us a better understanding in this field.

Variations in the central corneal thickness during the menstrual cycle in Indian women.

PURPOSE: To determine the changes in central corneal thickness (CCT) during the menstrual cycle in Indian women. METHODS: A prospective observational clinical study at a tertiary care center between December 2015 and December 2018. One hundred and twenty sixty women between 18 and 45 years were included. The CCT was measured using an ultrasound pachymeter at three specific timelines of the menstrual cycle: at the beginning (1st to 3rd day), during ovulation time (14th to 16th day), and at the end of the cycle (28th to 33rd day). Phases of the cycle were confirmed by the urine luteinizing hormone level. RESULTS: The mean CCT of both eyes was 541.76 ± 4.21 µm, 559.21 ± 4.50 µm, and 544.52 ± 8.06 µm at the beginning, mid, and end of cycle, respectively. The mean CCT of the right eye was 541.68 ± 4.15 µm, 559.08 ± 4.50 µm, and 544.44 ± 8.06 µm and of the left eye was 541.84 ± 4.27 µm, 559.35 ± 4.50 µm, and 544.61 ± 8.06 µm at the beginning, mid, and end of cycle, respectively. CONCLUSION: The CCT value was significantly (P < 0.001) higher during ovulation compared to the beginning and end of the menstrual cycle. Our study recommends adding menstrual history in the workup of women undergoing refractive surgery as physiological variations in the CCT may result in unexpected surgical outcomes.

Effect of Dickkopf-1 and colony stimulating factor-2 on the developmental competence, quality, gene expression and live birth rate of buffalo (Bubalus bubalis) embryos produced by hand-made cloning.

A functional canonical WNT signaling pathway exists in preimplantation embryos and inhibits embryonic development. Recent studies suggest that this pathway is over-expressed in nuclear transferred (NT), compared to IVF embryos. The present study investigated the effects of Dickkopf-1 (DKK1), an inhibitor of canonical WNT signaling pathway and colony stimulating factor-2 (CSF2), an embryokine, on the developmental competence, quality, gene expression and live birth rate of NT buffalo embryos produced by Hand-made cloning (HMC). Following supplementation of the in vitro culture medium on day 5 with DKK1 (100 ng/mL), CSF2 (10 ng/mL), DKK1+CSF2 or no supplementation (control), the blastocyst rate was higher (P < 0.05) with DKK1 and DKK1+CSF2 (42.6 ± 1.4% and 46.6 ± 0.9%, respectively) than with CSF2 or controls (40.6 ± 1.3% and 39.0 ± 1.3%, respectively). The apoptotic index of the blastocysts was lower (P < 0.05) for DKK1, CSF2 and DKK1+CSF2 groups (3.44 ± 0.14, 3.39 ± 0.11 and 3.11 ± 0.22, respectively) compared to controls (6.64 ± 0.25), and was similar to that of the IVF blastocysts (3.67 ± 0.18). Although the total cell number was similar for the DKK1, CSF2, DKK1+CSF2 and control groups (200.4 ± 3.05, 196.4 ± 3.73, 204.7 ± 3.71 and 205 ± 4.03, respectively), the inner cell mass:trophectoderm cell number ratio of DKK1, CSF2 and DKK1+CSF2 groups (0.21 ± 0.01, 0.17 ± 0.01 and 0.22 ± 0.02, respectively) was higher (P < 0.05) than controls (0.13 ± 0.01) and was similar to that of IVF blastocysts (0.19 ± 0.01). Treatment with DKK1 or CSF2 or both increased (P < 0.05) the expression level of OCT4, NANOG,SOX2, GATA6, BCL2, PTEN, P53, FGF4, GLUT1 and IFN-τ, and decreased that of C-MYC, CDX2, CASPASE, DNMT3a, TCF7 and LEF1 in blastocysts, compared to controls. Transfer of DKK1-treated embryos to 13 recipients resulted in 4 pregnancies (30.8%; 2 live births, one abortion and one currently at 9 months of pregnancy) whereas, transfer of DKK1+CSF2-treated embryos to 16 recipients, resulted in 4 pregnancies (25.0%), all of which resulted in live births. No pregnancy was obtained after transfer of control and CSF-treated embryos to 12 and 16 recipients, respectively. These results suggest that DKK1 treatment of NT embryos increases the blastocyst, conception and live birth rate, and improves their quality whereas, CSF2 treatment, does not affect the blastocyst, conception and live birth rate despite improvement in embryo quality.

DNA damage response proteins and its role in tumor progression of uveal melanoma with patient outcome.

BACKGROUND: The role of DNA damage response (DDR) proteins is poorly understood in uveal melanoma. ATR belongs to one of those proteins that induce DDR by arresting the cell cycle which leads to DNA repair. ATR is localized at position 23 on the same chromosome 3 where BAP1 is located at position 21.1 which is a known poor prognostic marker of UM. The aim of our study is to detect the expression of ATR at the protein and RNA levels and determine its prognostic significance. METHODS: Expression of nuclear ATR was investigated on sixty-nine UM patients. Formalin-fixed paraffin-embedded choroidal melanoma samples were taken to evaluate the expression of ATR. Fifty samples were also validated by real-time PCR. Results of both protein and mRNA were then correlated with clinicopathological parameters. To determine the prognostic significance, Kaplan-Meier and multivariate analyses were performed. RESULTS: Loss of ATR protein was seen in 72% cases which was statistically significant with epithelioid cell type (p = 0.005), tumor thickness (p = 0.016), mitotic figures (p = 0.001) and BAP1 loss (p < 0.001). At the transcriptional level loss of ATR was seen in 76% cases which were statistically significant with metastasis (p = 0.046), staging (0.044) and loss of BAP1 (p = 0.022). On multivariate analysis loss of ATR and tumor staging came out to be independent prognostic parameters. CONCLUSION: Our data suggest that ATR might serve as a potential prognostic marker in UM patients and could serve as a potential therapeutic target.

Constitutive expression of c-REL in uveal melanoma patients: correlation with clinicopathological parameters and patient outcome.

PURPOSE: Uveal melanoma (UM) is the most common intraocular cancer with a high mortality rate that requires new research in the field of prevention and treatment. c-REL is a member of the nuclear factor κB (NF-κB) transcription factor family and an emerging regulator of tumorigenesis. Therefore, the objective of the study is to evaluate the constitutive expression of c-REL in uveal melanoma patients and its prognostic significance. METHODS: Detection of c-REL expression was carried out by immunohistochemistry in all 75 patients, and qRT-PCR performed on 58 fresh cases of uveal melanoma along with IL-6 status. Immunoblot was performed to validate immunohistochemistry results. Expression of c-REL protein correlated with clinicopathological parameters and overall survival of patients. RESULTS: Immunohistochemistry results revealed nuclear expression of the c-REL protein (56%) in our cases. Out of 75 cases, 31 cases showed nuclear expression, and 11 cases had cytoplasmic expression. qRT-PCR showed upregulation of the REL gene in 56.89% cases at the transcriptional level. There was a statistically significant difference in the overall survival of patients with c-REL nuclear immunopositivity (p = 0.0048). On multivariate analysis, scleral invasion and c-REL nuclear expression found to be an independent prognostic factor (p < 0.05) CONCLUSIONS: To the best of our knowledge, this was the first study reporting the expression of the c-REL protein in uveal melanoma. Strong nuclear immunoexpression of c-Rel suggests NFκB pathway activation which might be involved in the progression of the disease. Differential expression of c-REL protein may be used as an attractive target for the development of anticancer strategies.

Successful transplantation of transfected enriched buffalo (Bubalus bubalis) spermatogonial stem cells to homologous recipients.

Genetic modification of spermatogonial stem cells (SSCs) is an alternative method to pronuclear microinjection and somatic cell nuclear transfer for transgenesis in large animals. In the present study, we optimized the process of homologous SSC transplantation in the water buffalo (Bubalus bubalis) using transfected enriched SSCs generated by a non-viral transfection approach. Firstly, the SSC enrichment efficiencies of extracellular matrix components viz. collagen, gelatin, and Datura stramonium agglutinin (DSA) lectin were determined either individually or in combination with Percoll density gradient centrifugation. The highest enrichment was achieved after differential plating with DSA lectin followed by Percoll density gradient centrifugation. Nucleofection showed greater transfection efficiency (68.55 ±â€¯4.56%, P < 0.05) for enriched SSCs in comparison to fugene HD (6.7 ±â€¯0.25%) and lipofectamine 3000 (15.57 ±â€¯0.74%). The transfected enriched SSCs were transplanted into buffalo males under the ultrasound guidance and testis was removed by castration after 7-8 weeks of transplantation. Persistence and localization of donor cells within recipient seminiferous tubules was confirmed using fluorescent microscopy. Further confirmation was done by flow cytometric evaluation of GFP expressing cells among those isolated from two-step enzymatic digestion of recipient testicular parenchyma. In conclusion, we demonstrated for the first time, generation of buffalo transfected enriched SSCs and their successful homologous transplantation in buffaloes. This study represents the first step towards genetic modifications in buffaloes using SSC transplantation technique.

Health risks from PAHs and potentially toxic elements in street dust of a coal mining area in India.

Polycyclic aromatic hydrocarbons (PAHs) and potentially toxic elements (PTEs) (Ba, Zn, Pb, Cu, Cr, Ni, As, Co) were determined in the road dusts of a coal mining area (Dhanbad, India) to assess their content and potential human health risks. Dust samples were collected from sign boards of the heavy traffic road connecting Dhanbad and Sindri. The total PAHs (∑PAHs, all values in mg/kg) content in the road dust samples varied from 3.98 to 13.1, with carcinogenic PAHs content of 14.8-34.4% of the ∑PAHs. Phenanthrene (2.72), fluorene (0.715) and pyrene (0.575) are the major PAHs. Principal component analysis revealed that these PAHs are probably originated from pyrogenic (coal combustion and traffic emission) and petrogenic (coal dust, tyre and road particles) sources. Among the PTEs, the mean content was higher for Ba (293 mg/kg) followed by Zn (224), Pb (128), Cu (52.6), Cr (45.2), Ni (22.0), As (17.5) and Co (8.11). The overall pollution load index varied from 0.43 to 1.0. Source analysis showed that PTEs in the road dust of the study site were derived from traffic emission (Zn, Fe, Mn, Co and Pb), coal dust (Cr, As and Ni) and soil (K, Mg, Ba, Sr and Ca). In general, the PTEs are lower, but the PAHs contents were elevated in the road dust samples. Although the exposure risks from PTEs are low, the risk to children (expressed as hazardous quotient) for As and Pb is near to the permissible limit of 1.0. Cancer risk from PAHs for adult (4.8 × 10-6) and child (5.3 × 10-6) has exceeded the acceptable limit of 10-6.

Calcium ionophore enhanced developmental competence and apoptotic dynamics of goat parthenogenetic embryos produced in vitro.

Parthenogenetically developed embryos are efficient sources of in vitro embryo production, having less ethical issue and being useful for investigating culture conditions/treatments, early developmental, genomic studies, and homonymous source of stem cells. Keeping its advantages in mind, we aimed to study the effects of different activating agents on embryo production and its quality and gene expression. In the present study, 1348 immature oocytes recovered were parthenogenetically developed to embryos. Usable-quality immature oocytes were collected by puncturing the surface follicles and matured in in vitro maturation (IVM) medium for 27 h in a humidified 5% CO2 incubator at 38.5°C. The matured oocytes were parthenogenetically activated by exposure to 5 µM calcium ionophore for 5 min or 7% ethanol for 7 min sequentially followed by 4 h incubation in 2 mM 6-DMAP and then in vitro cultured (IVC) in RVCL/G-2 medium for 8 days. Matured oocytes were activated by calcium ionophore, the cleavage rate observed was 76.67 ± 3.47%, and further they developed into 4-cell, 8-16-cell, morula, blastocyst, and hatched blastocyst with 85.30 ± 1.57%, 70.60 ± 2.00%, 45.05 ± 2.66%, 22.89 ± 2.40%, and 5.70 ± 1.97%, respectively. Whereas ethanol-activated oocytes showed cleavage rate of 87.60 ± 1.70% and further culture developed into 4-cell, 8-16 cell, morula, blastocyst, and hatched blastocyst with 86.14 ± 1.03%, 71.56 ± 2.21%, 40.90 ± 2.45%, 19.02 ± 1.26%, and 2.22 ± 0.38%, respectively. Blastocyst developed from calcium ionophore-activated oocytes showed significantly (P < 0.05) higher total cell number (282.25 ± 27.02 vs 206.00 ± 40.46) and a lower apoptotic index (2.42 ± 0.46 vs 4.07 ± 1.44) than blastocyst developed from ethanol-activated oocytes. The relative expression of anti-apoptotic genes (BCL2, BCL2A1, MCL) at different stages of embryos produced by either calcium ionophore or ethanol activation was found to be increased in earlier stages and decreased in later stages of embryonic development. Similarly, when these embryos were subjected to pro-apoptotic genes (BAX, BAD, BAK), expression was found to be slightly higher in blastocysts than other stages. This study shows that calcium ionophore-activated blastocysts were developmentally more competent than the ethanol-activated blastocysts.

Utility of a Single-Tube, Six-Color Flow Cytometry Panel for the Diagnosis of Myelodysplastic Syndrome: Experience of a Tertiary Care Centre in India.

Background: Diagnosis of myelodysplastic syndromes (MDS) is challenging in the presence of morphological mimickers. Flow cytometric immunophenotyping (FCI) has been added to the diagnostic armamentarium, but its use in clinical practice is variable. Materials and Methods: Bone marrow aspirate samples from 54 patients with a clinical and/or morphological suspicion of MDS were subjected to FCI using a single-tube, 6-colour panel comprising of monoclonal antibodies against CD13, CD11b, CD16, CD34, CD45 and CD56. Analysis was centered on the abnormal maturation pattern of granulocytes, blast percentage (≥3%) and ratio of side scatter peak channel value (SSC-PCV) of granulocytes and lymphocytes. Each of these parameters was assigned a score of 1. Overall sensitivity and specificity of this panel was ascertained to differentiate cytopenia/s of MDS from non-MDS cases. Results: Forty MDS and 14 non-MDS cases were diagnosed based on morphology and cytogenetic results. Twenty control samples were also processed simultaneously for FCI to assign the cutoff for various flow cytometric parameters. A score ≥2 was defined as positive for MDS. Hypogranularity was present in 62.5% cases of MDS. The median SSC-PCV of granulocytes and lymphocytes was 6.16 in the MDS group, 7.9 in the non-MDS group and 8.90 in the control group (p <0.05). This cut-off value of 6.16 had a specificity of 92.5% based on the ROC curve analysis. Abnormal granulocyte maturation patterns for CD13/16, CD13/11b and CD11b/16 dot plots were observed in 95.3, 69.8 and 74.4% cases, respectively. The overall sensitivity and specificity of the panel was found to be 87.5% and 64.2%, respectively. Conclusion: FCI is now an important tool for diagnostic workup in patients presenting with persistent cytopenia with or without morphological evidence of dyspoiesis. Inclusion of objective parameters like SSC-PCV would also reduce inter-lab variability in MDS diagnosis.

Does NEMO/IKKγ protein have a role in determining prognostic significance in uveal melanoma?

PURPOSE: Uveal melanoma, although a rare form of cancer, is the most common primary malignancy of the eye in adults. Nuclear factor-κB (NF-κB) is a transcription factor that transactivates genes involved in the regulation of cell growth, apoptosis, angiogenesis, and metastasis, but the molecular mechanisms that negatively regulate NF-κB activation are not fully understood. NF-κB can also be activated by DNA damage pathway through NEMO protein. Therefore, the objective of this study is to elucidate the role of NEMO/IKKγ protein in uveal melanoma patients. METHODS: Seventy-five formalin-fixed paraffin-embedded prospective tissues of uveal melanoma were included in the present study. These cases were reviewed and investigated for the expression of NEMO/IKKγ protein by immunohistochemistry and validated by western blotting along with the qRT-PCR for mRNA expression. Expression levels were correlated with the clinicopathological parameters and patients' outcome. RESULTS: Immunohistochemistry showed cytoplasmic expression of NEMO/IKKγ expression in only 22 out of 75 (29.33%) cases. This result was confirmed by western blotting, and correlated well with the immunohistochemical expression of NEMO/IKKγ protein (48 kDa). In addition, downregulation of this gene was found in 87.93% of the cases when compared with the normal tissues. On statistical analysis, loss of NEMO/IKKγ protein was correlated with neovascularization, high mitotic count, and presence of vascular loop (p < 0.05). There was less overall survival rate with low expression of NEMO/IKKγ protein in patients with uveal melanoma. CONCLUSION: This was the first study suggesting the relevant role of NEMO/IKKγ protein, and highlights the prognostic significance with outcome in uveal melanoma patients. This protein might be used as a screening biomarker in these patients after large-scale validation and translational studies.

The triple-negative (CD34-/HLA-DR-/CD11b-) profile rapidly and specifically identifies an acute promyelocytic leukemia.

INTRODUCTION: The genetic testing to confirm or rule out an acute promyelocytic leukemia (APL) typically takes a minimum of 24-72 hours. Flow cytometric immunophenotyping (FCI) on the other hand provides rapid and objective information to differentiate APL from non-APL. METHODS: FCI features, with single-tube 8-color combination using CD45, CD34, HAL-DR, CD11b, CD13, CD33, and CD117 and CD64, were compared for the 30 consecutive APL and 30 non-APL acute myeloid leukemia (AML) cases which morphologically mimicked an APL. The diagnosis was confirmed by cytogenetic or molecular genetic testing in the form of t (15:17) (q22; q21)/variant translocations or PML-RARA fusion transcript analysis. RESULTS: The APL cells lacked CD34, HLA-DR, and CD11b in 90%, 90%, and 93.3% cases, respectively. Myeloid antigens such as CD33, CD13, CD117, and CD64 were expressed in 96.7%, 96.7%, 76.7%, and 70% cases, respectively. The dual negative profiles, CD34-/HLA-DR- or HLA-DR-/CD11b-, were noted in 90% and 93.3% cases. The triple-negative (CD34-/HLA-DR-/CD11b-) profile was noted in 90% of the cases. The sensitivity, specificity, and positive predictive value (PPV) of CD34-/HLA-DR- and HLA-DR-/CD11b- profiles for the diagnosis of APL were found to be 90%, 80% & 81.1% and 93.3%, 86.7%& 87.5%, respectively. Combining the above two profiles resulted in a triple-negative profile (CD34-, HLA-DR- and CD11b-), which had a better specificity (93.3%) and positive predictive value (93.1%), with similar sensitivity. CONCLUSION: FCI is a rapid and reliable modality for the diagnosis of an APL. The triple-negative profile (CD34-/HLA-DR-/CD11b-) rapidly and specifically identifies an APL case.

Retro-bulbar cyst of Coenurus gaigeri in Barbari goats.

Two adult Barbari goats were presented to the Division of Animal Health of the Institute with the history of unilateral protrusion of the left eye ball, blepharitis and congestion of the conjunctival membrane. Physical and ultrasonographical examination revealed large fluctuating fluid filled bladder with distinct dimensions. The cysts were successfully removed along with its membrane and parasitological examination revealed it as a Coenurus gaigeri, the intermediate stage of T. multiceps gaigeri. The present report describes the retro-bulbar cyst of Coenurus gaigeri in two Barbari goats. This is the first report of retro-bulbar cyst of Coenurus gaigeri in goats.

Fluorescent Aerolysin (FLAER)-based paroxysmal nocturnal hemoglobinuria (PNH) screening: a single center experience from India.

BACKGROUND: Fluorescent aerolysin (FLAER) has been recommended as an important part of antibody panel used for flow cytometric detection of paroxysmal nocturnal hemoglobinuria (PNH) clone. This study was aimed to observe the frequency of PNH-positive clones and their sizes in patients screened for various indications. METHOD: A retrospective analysis of 624 patients screened over a period of 30 months. Frequency and size of clone sizes noted, and laboratory parameters were compared among different groups of patient being screened. RESULTS: There were 445 adults and 179 pediatric patients. Indications for screening included AA (n = 433), myelodysplastic syndrome (MDS) (n = 34), hemolytic anemia (n = 84), and thrombophilia workup group (n = 63). PNH clones were found in 39.03%, 5.88%, 26.19%, and 1.59% cases, respectively. No significant difference among adult or pediatric population was noted. The bone marrow failure (BMF) group [AA and MDS] with PNH clone had a significantly lower clone size (Median- 2.7%) as compared to classic PNH group (Median-77.2%). Most of the classic PNH cases (78.26%) and a small proportion of AA (9.9%) showed a large clone size (>50%). In spite of having large clone size, there was a significant difference between the median LDH values of these two groups (2511.5 vs 593 U/L). CONCLUSION: FLAER-based screening detects the presence of PNH clone in a high proportion of AA patients and some MDS patients. These patients usually have a small clone size. Even if they have a large clone, it does not get translated into a high LDH or severe clinical symptoms.

Role of CD200 in differential diagnosis of mature B-cell neoplasm.

INTRODUCTION: CD 200 is a type I immunoglobulin super family membrane glycoprotein, which is expressed in various mature B-cell neoplasm (MBN). This study aimed at analyzing the expression pattern of CD200 by flow cytometry immunophenotyping (FCI) and to evaluate its utility in narrowing down the differential diagnosis of MBN, particularly in low-grade lymphomas. METHODS: A total of 160 samples were evaluated by FCI over a period of 2 years (July 2014-June 2016), by a panel of antibodies including CD200. The mean fluorescence intensity (MFI) of CD200 in the neoplastic population was noted and compared among several groups of MBN. RESULTS: All the 98 cases of chronic lymphocytic leukemia (CLL), five being CD23-negative, expressed CD200 with moderate-to-bright intensity (median MFI: 1174). None of the 24 mantle cell lymphoma (MCL) cases, two being CD23-positive, expressed CD200 (median MFI: 10). All six hairy cell leukemia (HCL) cases expressed CD200. CD200 expression in HCL was brightest among all the MBNs, with a median MFI of 5050. Other MBN (n = 32) expressed CD200 in a proportion of cases and with variable intensity, usually dimmer than CLL. CONCLUSION: CD200 has a valid role in differentiating CLL from MCL, especially in the cases with immunophenotypic overlap. HCL has a consistent and brightest expression of CD200. It would be prudent to include CD200 in the primary panel of antibodies for MBN analysis.

Robotics in hepatobiliary surgery-initial experience, first reported case series from India.

INTRODUCTION: Robotic surgical system's ability to perform complex hepatobiliary surgeries is gaining momentum with outcomes similar to open surgery and advantages of minimal access surgery. The authors present their initial experience of a heterogenous spectrum of robotic hepatobiliary cases and the first reported case series from India. METHODS: Retrospective review of hepatobiliary cases done robotically from February 2015 to January 2016 was done. RESULTS: The series has ten patients; with median age of 45 years (range 15-72). Etiologies were choledochal cyst type IVa, benign lower end common bile duct stricture (biliary reconstruction group); incidental gallbladder carcinoma, hepatocellular carcinoma, recurrent pyogenic cholangitis, polycystic liver disease, hemangioma, liver metastases, hydatid cyst (resection group). Median operative duration was 510min; one patient needed intra-operative blood transfusion and there were no conversions to open surgery. One patient developed bile leak which was managed by biliary stenting and another thrombotic thrombocytopenic purpura during post-operative period. Median length of hospital stay was 6days with average cost of robotic surgery being $1700 USD more for major hepatectomy and $900 USD more for biliary reconstruction compared to open procedure. CONCLUSION: This initial series adds to existing data on the feasibility of robotic hepatobiliary cases with inherent advantages of minimal invasive surgery, however with limitation of availability and use of devices like cavitron ultrasonic surgical aspirator (CUSA) and higher operative cost.