Lysine 27 dimethylation of Drosophila linker histone dH1 contributes to heterochromatin organization independently of H3K9 methylation.

Post-translational modifications (PTMs) of core histones are important epigenetic determinants that correlate with functional chromatin states. However, despite multiple linker histone H1s PTMs have been identified, little is known about their genomic distribution and contribution to the epigenetic regulation of chromatin. Here, we address this question in Drosophila that encodes a single somatic linker histone, dH1. We previously reported that dH1 is dimethylated at K27 (dH1K27me2). Here, we show that dH1K27me2 is a major PTM of Drosophila heterochromatin. At mitosis, dH1K27me2 accumulates at pericentromeric heterochromatin, while, in interphase, it is also detected at intercalary heterochromatin. ChIPseq experiments show that >98% of dH1K27me2 enriched regions map to heterochromatic repetitive DNA elements, including transposable elements, simple DNA repeats and satellite DNAs. Moreover, expression of a mutated dH1K27A form, which impairs dH1K27me2, alters heterochromatin organization, upregulates expression of heterochromatic transposable elements and results in the accumulation of RNA:DNA hybrids (R-loops) in heterochromatin, without affecting H3K9 methylation and HP1a binding. The pattern of dH1K27me2 is H3K9 methylation independent, as it is equally detected in flies carrying a H3K9R mutation, and is not affected by depletion of Su(var)3-9, HP1a or Su(var)4-20. Altogether these results suggest that dH1K27me2 contributes to heterochromatin organization independently of H3K9 methylation.

Field performance and cost-effectiveness of a point-of-care triage test for HIV virological failure in Southern Africa.

INTRODUCTION: Antiretroviral therapy (ART) monitoring using viral load (VL) testing is challenging in high-burden, limited-resources settings. Chemokine IP-10 (interferon gamma-induced protein 10) strongly correlates with human immunodeficiency virus (HIV) VL. Its determination could serve to predict virological failure (VF) and to triage patients requiring VL testing. We assessed the field performance of a semi-quantitative IP-10 lateral flow assay (LFA) for VF screening in South Africa, and the cost-effectiveness of its implementation in Mozambique. METHODS: A cross-sectional study was conducted between June and December 2021 in three primary health clinics in the Western Cape. Finger prick capillary blood was collected from adults on ART for ≥1 year for direct application onto the IP-10 LFA (index test) and compared with a plasma VL result ≤1 month prior (reference test). We estimated the area under the receiver operating characteristic curves (AUC), sensitivity and specificity, to evaluate IP-10 LFA prediction of VF (VL>1000 copies/ml). A decision tree model was used to investigate the cost-effectiveness of integrating IP-10 LFA combined with VL testing into the current Mozambican ART monitoring strategy. Averted disability-adjusted life years (DALYs) and HIV acquisitions, and incremental cost-effectiveness ratios were estimated. RESULTS: Among 209 participants (median age 38 years and 84% female), 18% had VF. Median IP-10 LFA values were higher among individuals with VF compared to those without (24.0 vs. 14.6; p<0.001). The IP-10 LFA predicted VF with an AUC = 0.76 (95% confidence interval (CI) 0.67-0.85), 91.9% sensitivity (95% CI 78.1-98.3) and 35.1% specificity (95% CI 28.0-42.7). Integrating the IP-10 LFA in a setting with 20% VF prevalence and 61% VL testing coverage could save 13.0% of costs and avert 14.9% of DALYs and 55.7% new HIV acquisitions. Furthermore, its introduction was estimated to reduce the total number of routine VL tests required for ART monitoring by up to 68%. CONCLUSIONS: The IP-10 LFA is an effective VF triage test for routine ART monitoring. Combining a highly sensitive, low-cost IP-10 LFA-based screening with targeted VL confirmatory testing could result in significant healthcare quality improvements and cost savings in settings with limited access to VL testing.

Indocyanine Green (ICG)-Guided Identification of Hypermetabolic Pancreatic Nodules in Focal Congenital Hyperinsulinism: A Case Report in a 3-Month-Old Infant.

Indocyanine green (ICG)-guided near-infrared fluorescence has been recently adopted in pediatric surgery, although its use in the treatment of congenital hyperinsulinism has not been reported. We present a case of focal congenital hyperinsulinism in which ICG-navigation with ICG was used during surgical treatment. A 3-month-old infant was referred to our institution from a peripheral hospital for episodes of persistent hypoglycemia since birth, with no response to intravenous treatment with diazoxide, octreotide, or hydrochlorothiazide. An abdominal positron emission tomography-computed tomography scan showed a hypermetabolic nodule in the proximal portion of the body of the pancreas, compatible with focal congenital hyperinsulinism. A heterozygous mutation in the ABCC gene (Ala1516Glyfs*19) frameshift type inherited from the father was identified, which supported this diagnosis. Laparoscopy-assisted surgery was performed with ICG-guided near-infrared fluorescence, with intravenous injection of 16 mg ICG (2 mg/mg), which allowed localization of the focal lesion in the body of the pancreas. The lesion was resected with bipolar electrocautery and intraoperative histological study confirmed complete resection. Plasma glucose values normalized 6 hours after surgery and the patient was discharged 5 days later. In conclusion, the use of ICG in the treatment of congenital hyperinsulinism helps to identify hypermetabolic pancreatic nodules, decreasing the likelihood of incomplete resection.

Ischemic preconditioning of the graft for intestinal transplantation in rats.

To test the hypothesis that two modalities of IPC should decrease acute rejection and BT after SBTx in rats. Orthotopic allogenic SBTx was performed from Wistar to BN. IPC was performed by 2 ' and 5 ' superior mesenteric artery clamping, following 2-min and 5-min reperfusion before graft cooling and retrieving. Donor-recipient sets were randomly allocated to five groups: IPC2m4d, IPC2m7d, IPC5min7d, and the control groups for the two end points; ctrl4d and ctrl7d. IRI, rejection, and BT were assessed after four or seven days depending on the groups. Measured variables included: histology, leukocyte activation by tissue MPO determination, and proinflammatory cytokines (IL-b and TNF-α) to assess inflammatory response. Leukocyte activation was significantly reduced in IPC2m7d in comparison with Ctrl and IPC5min7d. Rejection tended to be lower in IPC2min7d. Cytokine levels were contradictory and not consistent with histology. Finally, BT was less frequent in IPC2min4d group but this benefit was missed in animals with rejection (7d). Inflammatory response (MPO) was reduced and rejection tended to be lower after in IPC2m7d. Bacterial translocation was reduced in IPC2min4d but the benefit was missed at day 7.

The use of sirolimus as a rescue therapy in pediatric intestinal transplant recipients.

To review our experience with SRL as a second-line therapy in our series of 45 SBTx recipients (1997-2009). Retrospective review of five children converted to SRL: 3 M/2 F; median of three yr old (range 20 months-18 yr); rescue indications, adverse events with SRL, resolution of tacrolimus-related side effects, incidence of rejection, PTLD, or GVHD were summarized. Tacrolimus was discontinued (average 13 months after transplant) because of refractory hemolytic anemia in four patients with decreased renal function and because of advanced renal failure and unclear neutropenia in one. PTLD and GVHD had been previously diagnosed in two. Tacrolimus-related side effects disappeared in all five although other immunosuppressants and splenectomy were used simultaneously or later in most of them. Adverse events reported after the conversion were infections (tuberculosis and Pneumocystis carinii in two) and mild hypertriglyceridemia. No rejection, GVHD, or PTLD episode was observed. Four patients are alive with excellent quality of life (median follow-up 18 months). Sirolimus is a safe rescue therapy in SBTx children when tacrolimus is not well tolerated. Renal function and hematologic disorders seem to improve, although other simultaneous strategies could be also involved. Further studies could demonstrate its efficacy as a first-line treatment.

The role of GLUT1 immunostaining in the diagnosis and classification of liver vascular tumors in children.

BACKGROUND/AIM: GLUT1 is an erythrocyte-type glucose transporter protein typically expressed in cutaneous proliferating hemangioma. Immunostaining for GLUT1 is becoming valuable for predicting the outcome of vascular skin tumors. Liver vascular tumors (LVTs) are a serious challenge for pediatric surgeons because of their often severe and sometimes unpredictable clinical course. To improve therapeutic strategies, we designed this study in which we tested in pathology specimens of LVT the hypothesis that GLUT1 expression could be useful to understand and classify LVT. MATERIAL AND METHODS: In the last 10 years, we treated 20 children with LVT. Pathology specimens from biopsy, excision, or autopsy were available in 11 of them. The paraffin sections were immunostained for GLUT1 and also for Ki-67 to assess the proportion of proliferating cells. Patients were divided into 2 groups: GLUT1-positive (n = 4) and GLUT1-negative (n = 7) that were compared for age at diagnosis, survival, and proportion of proliferating cells. Nonparametric and chi 2 tests were used for statistical analysis as appropriate. RESULTS: Mean age at diagnosis was higher in GLUT1-positive group, although not statistically significant in comparison with GLUT1-negative (308 +/- 515 vs 70 +/- 51 days, respectively). Three of 4 children in GLUT1-positive group died versus 1 of 7 in the GLUT1-negative group (not significant). All GLUT1-positive tumors were multicentric hemangiomata without central necrosis and only 1 with large vessels. Among GLUT1-negative tumors, 5 were solitary masses and 2 were multicentric (the value of the last 2 specimens was uncertain), 2 had central necrosis, and 2 had large vessels. Proliferation index was 18% +/- 1.42% and 1.42% +/- 0.97%, respectively, in each group ( P < .05). CONCLUSIONS: GLUT1-positive tumors have significantly higher proliferation rates than negative ones. Mortality tended to be higher in children with GLUT1-positive tumors. Positive GLUT1 immunostaining is likely specific for proliferating hemangioma, and it predicts the typical course of proliferation followed by involution.