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BACKGROUND AND OBJECTIVE: To determine how the gas concentration in air required to achieve full postoperative vitreous cavity fill varies in different aqueous outflow states. MATERIALS AND METHODS: A mathematical model was used to estimate gas dynamics. The change in gas bubble volume over time was calculated in an eye with normal aqueous outflow, ocular hypertension (OHT), and OHT with apraclonidine treatment. RESULTS: The concentration required was higher for all gases to achieve a full postoperative fill in OHT eyes versus normal eyes. Optimal gas concentrations were 22.6% for SF 6 , 13.9% for C 2 F 6 , and 11.6% for C 3 F 8 . Despite this, in OHT, the fill achieved was 95%, 95%, and 94% for SF 6 , C 2 F 6 , and C 3 F 8 , respectively. With apraclonidine, percentage fill improved for all gases. CONCLUSIONS: This is the first study to show aqueous outflow affects bubble size and indicates eyes with reduced outflow are at risk of underfill. This can ultimately affect surgical success. [ Ophthalmic Surg Lasers Imaging Retin a. 2020;51:522–528.]

Background Covid-19 significantly affected healthcare delivery over the past year, with a shift in focus away from nonurgent care. Emerging data are showing that screening for breast and colon cancer has dramatically decreased. It is unknown whether the same trend has affected patients with melanoma. Methods This is a retrospective cohort study of melanoma patients at two large-volume cancer centers. Patients were compared for 8 months before and after the lockdown. Outcomes focused on delay in treatment and possible resultant upstaging of melanoma. Results A total of 375 patients were treated pre-lockdown and 313 patients were treated post-lockdown (17% decrease). Fewer patients presented with in situ disease post-lockdown (15.3% vs. 17.9%), and a higher proportion presented with stage III-IV melanoma (11.2% vs. 9.9%). Comparing patients presenting 2 months before versus 2 months after the lockdown, there was an even more significant increase in Stage III-IV melanoma from 7.1% to 27.5% (p < 0.0001). Finally, in Stage IIIB-IIID patients, there was a decrease in patients receiving adjuvant therapy in the post lockdown period (20.0% vs. 15.2%). Conclusions As a result of the recent pandemic, it appears there has been a shift away from melanoma in situ and toward more advanced disease, which may have significant downstream effects on prognosis and could be due to a delay in screening. Significantly patients have presented after the lockdown, and fewer patients are undergoing the recommended adjuvant therapies. Patient outreach efforts are essential to ensure that patients continue to receive preventative medical care and screening as the pandemic continues.

TENB2, a transmembrane proteoglycan protein, is a promising target for antibody drug conjugate (ADC) therapy due to overexpression in human prostate tumors and rapid internalization. We previously characterized how predosing with parental anti-TENB2 monoclonal antibody (mAb) at 1 mg/kg in a patient-derived LuCap77 explant model with high (3+) TENB2 expression could (i) block target-mediated intestinal uptake of tracer (& 0.1 mg/kg) levels of radiolabeled anti-TENB2-monomethyl auristatin E ADC while preserving tumor uptake, and (ii) maintain efficacy relative to ADC alone. Here, we systematically revisit this strategy to evaluate the effects of predosing on tumor uptake and efficacy in LuCap96.1, a low TENB2-expressing (1+) patient-derived model that is more responsive to ADC therapy than LuCap77. Importantly, rather than using tracer (& 0.1 mg/kg) levels, radiolabeled ADC tumor uptake was assessed at 1 mg/kg – one of the doses evaluated in the tumor growth inhibition study – in an effort to bridge tissue distribution (PK) with efficacy (PD). Predosing with mAb up to 1 mg/kg had no effect on efficacy. These findings warrant further investigations to determine whether predosing prior to ADC therapy might improve therapeutic index by preventing ADC disposition and possible toxicological liabilities in antigen-expressing healthy tissues.

BACKGROUND/AIMS: Poor visual outcomes have recently been reported in black patients undergoing vitrectomy and delamination surgery for complications of proliferative diabetic retinopathy. We therefore investigated the outcome of surgery on a similar cohort of black patients and examined for potential predictors of visual success. METHODS: A single-centre retrospective case review of consecutive black patients who underwent vitrectomy and delamination surgery for complications of PDR between July 2010 and September 2017. The primary outcome measure was change in visual acuity (VA) at 6 months post-operatively. Multiple linear regression analysis was performed to evaluate determinants of change in VA. RESULTS: A total of 44 eyes of 44 patients were included. Mean age was 53.7 (range: 24.3–75.8) years. In all, 43% were male. A total of 52% had adjunctive pre-operative anti-VEGF therapy. Mean pre-operative VA was 1.49 logMAR ± 0.73 (range 0.18 to 2.6 logMAR). Mean change in VA at 6 months was a gain of 0.59 ± 0.94 logMAR (range 1.9 logMAR acuity loss to 2.5 logMAR acuity gain). Four cases required further surgery to treat rhegmatogenous detachment. The acuity of 68% improved by 0.3 or more logMAR. Silicone oil was used as primary tamponade in 7%. Pre-operative VA and use of silicone oil significantly predicted VA decline at 6 months (p = 0.001 and p = 0.007). CONCLUSIONS: The majority of our cohort derived visual benefit from vitrectomy and delamination for PDR-related complications. Improvement in VA was comparable to outcomes from the UK National Ophthalmic Database report. Silicone oil should be avoided these patients if possible.

BACKGROUND: Ophthalmic emergencies comprise a significant portion of junior doctors’ workload in accident and emergency (A&E). However, previous studies have demonstrated a lack of training and confidence in the management of such emergencies. This study assessed changes in basic ophthalmic training that A&E junior doctors received in dealing with eye emergencies, their perceived level of confidence and the availability of appropriate ophthalmic equipment in A&E over the last 15 years. METHODS: A prospective, national, combined online and telephone survey using a previously published questionnaire was performed. Foundation year two doctors (FY2s) from each A&E department in the UK listed on the official NHS directory were contacted for participation. RESULTS: Two hundred and ten A&E departments were contacted and 202 responded (response rate of 96.2%). There was no significant change in the number of A&E departments equipped with slit lamps (82.5% in 2003 vs 79.7% in 2018; p = 0.26). However, the prevalence of training in its use has decreased significantly (68.4% in 2003 vs 52% in 2018; p = 0.005). There was also a significant reduction in the prevalence of training in the management of eye emergencies (77.4% in 2003 vs 45.5% in 2018; p

AIMS: To report anatomical and functional outcomes in patients with proliferative sickle retinopathy (PSR) who underwent 23-gauge (23G) and 20-gauge (20G) vitrectomy. METHODS: Retrospective consecutive case series of patients who underwent vitreoretinal intervention for complications of PSR between April 2009 and February 2015. Operations were performed at a tertiary referral centre, Moorfields Eye Hospital. Visual acuity and anatomical success rates were evaluated for PSR complicated by retinal detachment, tractional vitreous haemorrhage and macular hole. Proliferative diabetic retinopathy cases were excluded. RESULTS: A total of 71 eyes (63 patients) underwent vitreoretinal surgery for PSR complications with 26 months mean follow-up. Primary indications were: tractional retinal detachment (TRD, n = 17), TRD with rhegmatogenous retinal detachment (n = 16), rhegmatogenous retinal detachment (n = 5, macula-on: 1, macula-off: 4), vitreous haemorrhage (n = 19), epiretinal membrane (n = 6), and full thickness macula hole (n = 8). Thirty-nine cases underwent 20G vitrectomy, and 23G surgery was performed in 32 eyes. Mean best corrected visual acuity (BCVA) improved from pre-operative 1.30 LogMAR to final BCVA of 0.74 LogMAR (p

PURPOSE To evaluate the real world clinical outcomes of intravitreal ocriplasmin in patients with vitreomacular traction (VMT) with and without full thickness macular holes (FTMH) treated according to NICE guidance. METHODS Retrospective observational case series of 25 patients treated with a single intravitreal ocriplasmin injection between December 2013 and December 2015. Best corrected visual acuity and optical coherence tomography exams were performed to determine visual outcomes and anatomical VMT release and FTMH closure over time. Two patient groups were identified: ocular macular co-morbidity (OCM) and no OCM (nOCM), with follow-up at 4, 12, and 24 weeks. RESULTS Twenty-five patients were identified that included 19 patients with VMT, and 6 patients with VMT plus FTMH. In the nOCM group of 22 patients, the release rate of VMT was 44%, 63%, and 69% at 4, 12 and 24 weeks respectively. In the "real-world" OCM group of 25 patients, the VMT release rate was 37%, 53%, and 58% at the same time-points. In both groups, the FTMH closure rate was 33%, 50%, and 67% at 4, 12, and 24 weeks. At mean follow-up of 30 weeks in the VMT group with nOCM, the mean LogMAR VA improved significantly from 0.44 to 0.28 (p = 0.0068, paired t-test). Three were no serious adverse events. CONCLUSIONS This study reports improved efficacy of intravitreal ocriplasmin for both VMT and FTMH, and is more favourable in patients with no ocular co-morbidity. We highlight the importance of careful patient selection and structured standard of care pathways to identify patients who will benefit from the positive visual and anatomical effects of intravitreal ocriplasmin.

For years, clinical and basic researchers have been aware of the presence of PTEN in the nucleus in cell culture, animal models, and both healthy and diseased human tissues. Despite the early recognition of nuclear PTEN, the understanding of the mechanisms of its nuclear localization, function in the nucleus, and importance in biology and human disease has been lacking. Over the last decade, emerging concepts for the complex involvement of nuclear PTEN in a variety of processes, including genome maintenance and DNA repair, cell-cycle control, gene expression, and DNA replication, are illuminating what could prove to be the key path toward a full understanding of PTEN function in health and disease. Dysregulation of nuclear PTEN is now considered an important aspect of the etiology of many pathologic conditions, prompting reconsideration of the therapeutic approaches aimed at countering the consequences of PTEN deficiency. This new knowledge is fueling the development of innovative therapeutic modalities for a broad spectrum of human conditions, from cancer and metabolic diseases, to neurological disorders and autism.

Objective Chronic hepatitis B virus (HBV) infection and liver cancer mortality represent a neglected health disparity among Asian Americans. The purposes of this study were to compare the prevalence of hepatitis B among a diverse group of 1311 Ohioans in Franklin County, OH (85% Asians, 7.5% African Americans, and 6.5% whites) and to improve access to care for high-risk Asian adults through advocacy and policy changes. Research Design and Methods The Asian subgroups comprised of Chinese, Filipino, Asian Indian, Pakistani, Vietnamese, Korean, Laotian, Indonesian, Japanese, Cambodian, Thai, and Malaysian nationalities. The HBV screening was completed at health fairs, restaurants, churches, and temples from 2006 to 2011. Results The prevalence of HBV infection (9.5% vs 5%) and family history of liver cancer was significantly higher among Asians than other racial ethnic groups ( P = 0.001). Cambodian, Vietnamese, and Chinese participants were disproportionately infected with the virus compared with other Asian subgroups ( P < 0.001). Advocacy and policy changes for resources allowed vaccine-eligible Asians included as “high risk” group to receive free vaccinations at the health department. However, although vouchers were provided to vaccine-eligible Asian adults, compliance in getting vaccinated was very low (11%). Common barriers for compliance were lack of time and knowledge of completing the 3 shot series, low English proficiency, and fear of adverse effects. Conclusions Outreach education may use community liaisons to improve screenings, education, and vaccination/treatment. A hepatitis free clinic was established in 2009 to provide culturally and linguistically appropriate treatment for low-income Asian Americans in Franklin County, OH.

Terminal differentiation of B cells and hypergammaglobulinemia are hallmarks of B-cell hyperactivity in HIV disease. Plasmablasts are terminally differentiating B cells that circulate transiently in the blood following infection or vaccination; however, in HIV infection, they arise early and are maintained at abnormally high levels in viremic individuals. Here we show that only a small fraction of plasmablasts in the blood of viremic individuals is HIV specific. Assessment of plasmablast immunoglobulin isotype distribution revealed increased IgG + plasmablasts in early and most prominently during chronic HIV viremia, contrasting with a predominantly IgA + plasmablast profile in HIV-negative individuals or in aviremic HIV-infected individuals on treatment. Of note, IgG is the predominant immunoglobulin isotype of plasmablasts that arise transiently in the blood following parenteral immunization. Serum immunoglobulin levels were also elevated in HIV-infected viremic individuals, especially IgG, and correlated with levels of IgG + plasmablasts. Several soluble factors associated with immune activation were also increased in the sera of HIV-infected individuals, especially in viremic individuals, and correlated with serum immunoglobulin levels, particularly IgG. Thus, our data suggest that while plasmablasts in the blood may contribute to the HIV-specific immune response, the majority of these cells are not HIV specific and arise early, likely from indirect immune-activating effects of HIV replication, and reflect over time the effects of chronic antigenic stimulation. Such B-cell dysregulation may help explain why the antibody response is inadequate in HIV-infected individuals, even during early infection.

Cancer sequencing predicts driver genes using recurrent protein-altering mutations, but detecting recurrence for non-coding mutations remains unsolved. Here, we present a convergence framework for recurrence analysis of non-coding mutations using three-dimensional co-localization of epigenomically-identified regions. We define the regulatory plexus of each gene as its cell-type-specific three-dimensional gene-regulatory neighborhood, inferred using Hi-C chromosomal interactions and chromatin state annotations. Using 16 matched tumor-normal prostate transcriptomes, we predict tumor-upregulated genes, and find enriched plexus mutations in distal regulatory regions normally repressed in prostate, suggesting out-of-context de-repression. Using 55 matched tumor-normal prostate genomes, we predict 15 driver genes by convergence of dispersed, low-frequency mutations into high-frequency dysregulation events along prostate-specific plexi, while controlling for mutational heterogeneity across regions, chromatin states, and patients. These putative drivers play roles in growth signaling, immune evasion, mitochondrial function, and vascularization, suggesting higher-order pathway-level convergence. We experimentally validate the PLCB4 plexus and its ability to affect the canonical PI3K cancer pathway.

Despite the rare appearance of potent HIV-neutralizing mAbs in infected individuals requiring prolonged affinity maturation, little is known regarding this process in the majority of viremic individuals. HIV-infected individuals with chronic HIV viremia have elevated numbers of nonconventional tissue-like memory (TLM) B cells that predominate in blood over conventional resting memory (RM) B cells. Accordingly, we investigated affinity maturation in these 2 memory B cell populations. Analysis of IgG-expressing TLM B cells revealed a higher number of cell divisions compared with RM B cells; however, TLM B cells paradoxically displayed significantly lower frequencies of somatic hypermutation (SHM). To assess Ab reactivity in TLM and RM B cells, single-cell cloning was performed on HIV envelope CD4–binding site–sorted (CD4bs-sorted) B cells from 3 individuals with chronic HIV viremia. Several clonal families were present among the 127 cloned recombinant mAbs, with evidence of crosstalk between TLM and RM B cell populations that was largely restricted to non-VH4 families. Despite evidence of common origins, SHM frequencies were significantly decreased in TLM-derived mAbs compared with SHM frequencies in RM-derived mAbs. However, both cell populations had lower frequencies of SHMs than did broadly neutralizing CD4bs–specific mAbs. There was a significant correlation between SHM frequencies and the HIV-neutralizing capacities of the mAbs. Furthermore, HIV neutralization was significantly higher in the RM-derived mAbs compared with that seen in the TLM-derived mAbs, and both SHM frequencies and neutralizing capacity were lowest in TLM-derived mAbs with high polyreactivity. Thus, deficiencies in memory B cells that arise during chronic HIV viremia provide insight into the inadequacy of the Ab response in viremic individuals.

Both human epidermal growth factor receptor 2 (HER-2/neu) and VEGF overexpression correlate with aggressive phenotypes and decreased survival among breast cancer patients. Concordantly, the combination of trastuzumab (anti-HER2) with bevacizumab (anti-VEGF) has shown promising results in preclinical xenograft studies and in clinical trials. However, despite the known antiangiogenic mechanism of anti-VEGF antibodies, relatively little is known about their effects on the pharmacokinetics and tissue distribution of other antibodies. This study aimed to measure the disposition properties, with a particular emphasis on tumor uptake, of trastuzumab in the presence or absence of anti-VEGF. Radiolabeled trastuzumab was administered alone or in combination with an anti-VEGF antibody to mice bearing HER2-expressing KPL-4 breast cancer xenografts. Biodistribution, autoradiography, and single-photon emission computed tomography–X-ray computed tomography imaging all showed that anti-VEGF administration reduced accumulation of trastuzumab in tumors despite comparable blood exposures and similar distributions in most other tissues. A similar trend was also observed for an isotype-matched IgG with no affinity for HER2, showing reduced vascular permeability to macromolecules. Reduced tumor blood flow (P < 0.05) was observed following anti-VEGF treatment, with no significant differences in the other physiologic parameters measured despite immunohistochemical evidence of reduced vascular density. In conclusion, anti-VEGF preadministration decreased tumor uptake of trastuzumab, and this phenomenon was mechanistically attributed to reduced vascular permeability and blood perfusion. These findings may ultimately help inform dosing strategies to achieve improved clinical outcomes. Mol Cancer Ther; 11(3); 752–62. ©2012 AACR.

Characterization of lymphocytes including B cells during early versus chronic HIV infection is important for understanding the impact of chronic viremia on immune cell function. In this setting, we investigated B cells before and after reduction of HIV plasma viremia by antiretroviral therapy (ART). At baseline, peripheral blood B-cell counts were significantly lower in both early and chronic HIV-infected individuals compared with uninfected controls. Similar to CD4+ but not CD8+ T cells, B-cell numbers in both groups increased significantly after ART. At baseline, B cells of early HIV-infected individuals were composed of a higher percentage of plasmablasts and resting memory B cells compared with chronic HIV-infected individuals whose B cells were composed of a higher percentage of immature/transitional and exhausted B cells compared with their early infection counterparts. At 1 year after ART, the percentage of resting memory B cells remained higher in early compared with chronic HIV-infected individuals. This difference translated into a better functional profile in that memory B-cell responses to HIV and non-HIV antigens were superior in early- compared with chronic-treated HIV infected individuals. These findings provide new insights on B cells in HIV infection and how early initiation of ART may prevent irreversible immune system damage.

Despite the demonstrated constitutive activation of NF-kappaB in nasopharyngeal carcinoma (NPC), the therapeutic potential of targeting this pathway has not been investigated. Here, we employed a small molecule inhibitor of NF-kappaB, DHMEQ (which mainly blocks nuclear translocation of activated NF-kappaB) and demonstrated significant inhibition of NPC cell proliferation, migration, invasion, as well as anchorage-independent growth. These antitumor effects were associated with induction of G(2)/M cell cycle arrest and apoptosis, and downregulation of NF-kappaB target genes (EGFR, cyclin D1 and survivin). This first demonstration of therapeutic benefits of NF-kappaB targeting in NPC implicates the importance of targeting this pathway in NPC.

Background. Untreated human immunodeficiency virus (HIV) disease leads to abnormalities in all major lymphocyte populations, including CD4 + T cells, CD8 + T cells, and B cells. However, little is known regarding the effect of antiretroviral therapy (ART)-induced decrease in HIV viremia on B cell numbers and subpopulations. Methods. We conducted a longitudinal study to evaluate changes in B cell numbers and subpopulations that occur during the course of 12 months of effective ART in a group of individuals with chronic HIV infection. Results. ART-induced decrease in HIV viremia was associated with a significant increase in B cell counts, similar to increases in CD4 + T cell counts yet distinct from the lack of increase in CD8 + T cells. The increase in B cell counts was accompanied by a significant decrease in the frequency of apoptosis-prone B cell subpopulations, namely mature activated and immature transitional B cells, which are overrepresented in untreated HIV disease. The increase in B cell counts was reflected by a significant increase in naive and resting memory B cells, both of which represent populations that are essential for generating adequate humoral immunity. Conclusions. Normalization of B cell counts and subpopulations may help to explain the improvement in humoral immunity reported to occur after an ART-induced decrease in HIV viremia.

Background We previously reported abnormalities in circulating B cells in patients with chronic granulomatous disease (CGD) and those with HIV infection. Gastrointestinal complications are common to both diseases and likely involve perturbation of immune cells, including plasma cells (PCs). IgA is the most abundant immunoglobulin in the human body, with roles in protection and maintenance of intestinal homeostasis. IgA is produced primarily by PCs residing in mucosal tissues that are also thought to circulate in the blood. Objective We sought to characterize and compare PCs in patients with infectious (HIV) and noninfectious (CGD and Crohn disease) diseases that have been associated with intestinal inflammation. Methods Phenotypic and transcriptional analyses were performed on cells isolated from the blood and colon. Results IgA-secreting CCR10-expressing PCs predominated in the guts of healthy subjects, whereas in patients with HIV, CGD, and Crohn disease, there was a significant increase in the proportion of IgG-secreting PCs. Where intestinal inflammation was present, IgG-secreting PCs expressed reduced levels of CCR10 and increased levels of CXCR4. The intensity of CXCR4 expression correlated with the frequency of IgG-expressing PCs and the frequency of CXCR4 + /IgG + PCs was associated with the severity of intestinal inflammatory disease yet distinct from PCs and plasmablasts circulating in the blood. Conclusions These findings suggest that regardless of the underlying disease, the presence of CXCR4 + /IgG + PCs in the gut is a strong yet localized indicator of intestinal inflammation. Furthermore, our findings suggest that CXCR4 + /IgG + PCs might play a role in immune cell homeostasis during inflammatory processes of the gut.

The practice of medicine is undergoing continual change. For trainees, the transition from a Halstedian apprenticeship to competence-based models of surgical training and assessment has been accelerated by European Working Time Directives, public/regulatory expectations and funding pressures. For qualified practitioners, revalidation will demand evidence of practice to a good standard. To successfully revalidate, ophthalmologists are expected to demonstrate competence if they undertake (phacoemulsification) cataract surgery, the commonest operation in the UK1. We pose two key questions: what are reasonable minima for (1) the number of operations before trainees can operate independently in a safe and timely manner (acquisition of competency) and (2) for consultants to maintain competency? Like our colleagues in other specialties, the Ophthalmic Specialty Training syllabus parallels the UK Intercollegiate Surgical Curriculum Programme, with objective checklists and global rating scales that qualitatively assess operative skills. One example is the Objective Structured Assessment of Technical skills, originally developed by general surgeons and now validated across various surgical disciplines including ophthalmology. Our focus on operative numbers is due to the delicate 4 μm window of error that phacoemulsification surgeons must contend with. Furthermore, in other specialties (e.g. ENT, plastics, maxillofacial surgery) microsurgery is deemed a higher surgical, non-core skill, whereas ophthalmic trainees are assessed from the very first year. To achieve Certificate of Completion of Training, plastic surgery trainees must perform at least 22 microvascular anastomoses. In comparison, the seven-year UK ophthalmology training programme specifies at least 350 full cataract operations. Three-year US residencies require 86 and in Australia ‘50 intraocular procedures’ are expected after two years of a five-year traineeship. Actual (median) cumulative totals are 500–600 for the UK2 versus 100 for the USA.3 With moves towards competence-based curricula, the intensity of hands-on surgical training may have a bearing on how rapidly trainees acquire operating skills, which is more relevant in US residencies where trainees usually gain surgical experience in their senior years. Trainees improve significantly to approach the benchmark of a published ophthalmologist's (longest) average operating time of 26.8 min after the 120th case.4 Posterior capsular rupture (PCR) is the commonest complication of phacoemulsification. Widely used as a surgical quality benchmark; it is a likely key quality indicator for revalidation.1 PCR rates range from >5% for junior trainees5 to

CD27+ memory B cells are reduced in the blood of patients with chronic granulomatous disease (CGD) for reasons and consequences that remain unclear. Here we confirm not only decreased CD27+ but also IgG+ B cells in the blood of CGD patients compared with healthy donors (HDs). However, among IgG+ B cells, the ratio of CD27− to CD27+ was significantly higher in CGD patients compared with HDs. Similar to conventional memory B cells, CD27−IgG+ B cells of CGD patients expressed activation markers and had undergone somatic hypermutation, albeit at levels lower than their CD27+ counterparts. Functional analyses revealed slight reductions in frequencies of total IgG but not influenza-specific memory B-cell responses, as measured by Elispot in CGD patients compared with HDs. Serum IgG levels and influenza-specific antibodies were also normal in these CGD patients. Finally, we provide evidence that influenza-specific memory B cells can be present within the CD27−IgG+ B-cell compartment. Together, these findings show that, despite reduced circulating CD27+ memory B cells, CGD patients maintain an intact humoral immunologic memory, with potential contribution from CD27− B cells.

Mendelian analysis of disorders of immune regulation can provide insight into molecular pathways associated with host defense and immune tolerance.We identified three families with a dominantly inherited complex of cold-induced urticaria, antibody deficiency, and susceptibility to infection and autoimmunity. Immunophenotyping methods included flow cytometry, analysis of serum immunoglobulins and autoantibodies, lymphocyte stimulation, and enzymatic assays. Genetic studies included linkage analysis, targeted Sanger sequencing, and next-generation whole-genome sequencing.Cold urticaria occurred in all affected subjects. Other, variable manifestations included atopy, granulomatous rash, autoimmune thyroiditis, the presence of antinuclear antibodies, sinopulmonary infections, and common variable immunodeficiency. Levels of serum IgM and IgA and circulating natural killer cells and class-switched memory B cells were reduced. Linkage analysis showed a 7-Mb candidate interval on chromosome 16q in one family, overlapping by 3.5 Mb a disease-associated haplotype in a smaller family. This interval includes PLCG2, encoding phospholipase Cγ(2) (PLCγ(2)), a signaling molecule expressed in B cells, natural killer cells, and mast cells. Sequencing of complementary DNA revealed heterozygous transcripts lacking exon 19 in two families and lacking exons 20 through 22 in a third family. Genomic sequencing identified three distinct in-frame deletions that cosegregated with disease. These deletions, located within a region encoding an autoinhibitory domain, result in protein products with constitutive phospholipase activity. PLCG2-expressing cells had diminished cellular signaling at 37°C but enhanced signaling at subphysiologic temperatures.Genomic deletions in PLCG2 cause gain of PLCγ(2) function, leading to signaling abnormalities in multiple leukocyte subsets and a phenotype encompassing both excessive and deficient immune function. (Funded by the National Institutes of Health Intramural Research Programs and others.).

Stimulation through TLR represents a new therapeutic approach for enhancing Ab responses to vaccination. Considering that Ab responses are decreased in HIV disease and that B cells express TLR9 and respond to TLR9 agonists, we investigated the responsiveness of B cell subpopulations from HIV-infected and uninfected individuals to the TLR9 agonist CpG oligonucleotide type B (CpG-B) in the presence and absence of BCR ligation and T cell help (CD40L). CpG-B was equally effective in stimulating the proliferation of naive B cells of HIV-infected individuals and HIV-negative individuals, and, when combined with BCR and CD40 ligation, cytokine secretion by naive B cells was also comparable in HIV-infected and uninfected individuals. In contrast, CD27+ memory/activated B cells of HIV-infected individuals with active disease were less responsive to CpG-B in terms of proliferation and cytokine secretion when compared with CD27+ B cells of HIV-negative and HIV-infected individuals whose viremia was controlled by antiretroviral therapy. These findings suggest that despite abnormalities in memory B cells of HIV-infected individuals with active disease, naive B cells of HIV-infected individuals, irrespective of disease status, can respond to TLR9 agonists and that the incorporation of such agents in vaccine formulations may enhance their Ab responses to vaccination.