Thyrotropin-releasing hormone (TRH) promotes wound re-epithelialisation in frog and human skin

There remains a critical need for new therapeutics that promote wound healing in patients suffering from chronic skin wounds. This is, in part, due to a shortage of simple, physiologically and clinically relevant test systems for investigating candidate agents. The skin of amphibians possesses a remarkable regenerative capacity, which remains insufficiently explored for clinical purposes. Combining comparative biology with a translational medicine approach, we report the development and application of a simple ex vivo frog (Xenopus tropicalis) skin organ culture system that permits exploration of the effects of amphibian skin-derived agents on re-epithelialisation in both frog and human skin. Using this amphibian model, we identify thyrotropin-releasing hormone (TRH) as a novel stimulant of epidermal regeneration. Moving to a complementary human ex vivo wounded skin assay, we demonstrate that the effects of TRH are conserved across the amphibian-mammalian divide: TRH stimulates wound closure and formation of neo-epidermis in organ-cultured human skin, accompanied by increased keratinocyte proliferation and wound healing-associated differentiation (cytokeratin 6 expression). Thus, TRH represents a novel, clinically relevant neuroendocrine wound repair promoter that deserves further exploration. These complementary frog and human skin ex vivo assays encourage a comparative biology approach in future wound healing research so as to facilitate the rapid identification and preclinical testing of novel, evolutionarily conserved, and clinically relevant wound healing promoters

Characteristics of the Earliest Cross-Neutralizing Antibody Response to HIV-1

Recent cross-sectional analyses of HIV-1+ plasmas have indicated that broadly cross-reactive neutralizing antibody responses are developed by 10%–30% of HIV-1+ subjects. The timing of the initial development of such anti-viral responses is unknown. It is also unknown whether the emergence of these responses coincides with the appearance of antibody specificities to a single or multiple regions of the viral envelope glycoprotein (Env). Here we analyzed the cross-neutralizing antibody responses in longitudinal plasmas collected soon after and up to seven years after HIV-1 infection. We find that anti-HIV-1 cross-neutralizing antibody responses first become evident on average at 2.5 years and, in rare cases, as early as 1 year following infection. If cross-neutralizing antibody responses do not develop during the first 2–3 years of infection, they most likely will not do so subsequently. Our results indicate a potential link between the development of cross-neutralizing antibody responses and specific activation markers on T cells, and with plasma viremia levels. The earliest cross-neutralizing antibody response targets a limited number of Env regions, primarily the CD4-binding site and epitopes that are not present on monomeric Env, but on the virion-associated trimeric Env form. In contrast, the neutralizing activities of plasmas from subjects that did not develop cross-neutralizing antibody responses target epitopes on monomeric gp120 other than the CD4-BS. Our study provides information that is not only relevant to better understanding the interaction of the human immune system with HIV but may guide the development of effective immunization protocols. Since antibodies to complex epitopes that are present on the virion-associated envelope spike appear to be key components of earliest cross-neutralizing activities of HIV-1+ plasmas, then emphasis should be made to elicit similar antibodies by vaccination

Profiles of Human Serum Antibody Responses Elicited by Three Leading HIV Vaccines Focusing on the Induction of Env-Specific Antibodies

In the current report, we compared the specificities of antibody responses in sera from volunteers enrolled in three US NIH-supported HIV vaccine trials using different immunization regimens. HIV-1 Env-specific binding antibody, neutralizing antibody, antibody-dependent cell-mediated cytotoxicity (ADCC), and profiles of antibody specificity were analyzed for human immune sera collected from vaccinees enrolled in the NIH HIV Vaccine Trial Network (HVTN) Study #041 (recombinant protein alone), HVTN Study #203 (poxviral vector prime-protein boost), and the DP6-001 study (DNA prime-protein boost). Vaccinees from HVTN Study #041 had the highest neutralizing antibody activities against the sensitive virus along with the highest binding antibody responses, particularly those directed toward the V3 loop. DP6-001 sera showed a higher frequency of positive neutralizing antibody activities against more resistant viral isolate with a significantly higher CD4 binding site (CD4bs) antibody response compared to both HVTN studies #041 and #203. No differences were found in CD4-induced (CD4i) antibody responses, ADCC activity, or complement activation by Env-specific antibody among these sera. Given recent renewed interest in realizing the importance of antibody responses for next generation HIV vaccine development, different antibody profiles shown in the current report, based on the analysis of a wide range of antibody parameters, provide critical biomarker information for the selection of HIV vaccines for more advanced human studies and, in particular, those that can elicit antibodies targeting conformational-sensitive and functionally conserved epitopes

The use of a bayesian hierarchy to develop and validate a co-morbidity score to predict mortality for linked primary and secondary care data from the NHS in England

Background: We have assessed whether the linkage between routine primary and secondary care records provided an opportunity to develop an improved population based co-morbidity score with the combined information on co-morbidities from both health care settings. Methods: We extracted all people older than 20 years at the start of 2005 within the linkage between the Hospital Episodes Statistics, Clinical Practice Research Datalink, and Office for National Statistics death register in England. A random 50% sample was used to identify relevant diagnostic codes using a Bayesian hierarchy to share information between similar Read and ICD 10 code groupings. Internal validation of the score was performed in the remaining 50% and discrimination was assessed using Harrell’s C statistic. Comparisons were made over time, age, and consultation rate with the Charlson and Elixhauser indexes. Results: 657,264 people were followed up from the 1st January 2005. 98 groupings of codes were derived from the Bayesian hierarchy, and 37 had an adjusted weighting of greater than zero in the Cox proportional hazards model. 11 of these groupings had a different weighting dependent on whether they were coded from hospital or primary care. The C statistic reduced from 0.88 (95% confidence interval 0.88–0.88) in the first year of follow up, to 0.85 (0.85–0.85) including all 5 years. When we stratified the linked score by consultation rate the association with mortality remained consistent, but there was a significant interaction with age, with improved discrimination and fit in those under 50 years old (C=0.85, 0.83–0.87) compared to the Charlson (C=0.79, 0.77–0.82) or Elixhauser index (C=0.81, 0.79–0.83). Conclusions: The use of linked population based primary and secondary care data developed a co-morbidity score that had improved discrimination, particularly in younger age groups, and had a greater effect when adjusting for co-morbidity than existing scores

Recombinant HIV Envelope Proteins Fail to Engage Germline Versions of Anti-CD4bs bNAbs

Vaccine candidates for HIV-1 so far have not been able to elicit broadly neutralizing antibodies (bNAbs) although they express the epitopes recognized by bNAbs to the HIV envelope glycoprotein (Env). To understand whether and how Env immunogens interact with the predicted germline versions of known bNAbs, we screened a large panel (N:56) of recombinant Envs (from clades A, B and C) for binding to the germline predecessors of the broadly neutralizing anti-CD4 binding site antibodies b12, NIH45-46 and 3BNC60. Although the mature antibodies reacted with diverse Envs, the corresponding germline antibodies did not display Env-reactivity. Experiments conducted with engineered chimeric antibodies combining the mature and germline heavy and light chains, respectively and vice-versa, revealed that both antibody chains are important for the known cross-reactivity of these antibodies. Our results also indicate that in order for b12 to display its broad cross-reactivity, multiple somatic mutations within its VH region are required. A consequence of the failure of the germline b12 to bind recombinant soluble Env is that Env-induced B-cell activation through the germline b12 BCR does not take place. Our study provides a new explanation for the difficulties in eliciting bNAbs with recombinant soluble Env immunogens. Our study also highlights the need for intense efforts to identify rare naturally occurring or engineered Envs that may engage the germline BCR versions of bNAbs

Regular use of analgesics is a risk factor for renal cell carcinoma

Phenacetin-based analgesics have been linked to the development of renal pelvis cancer and renal cell carcinoma (RCC). The relationship between non-phenacetin types of analgesics and kidney cancer is less clear, although laboratory evidence suggests that these drugs possess carcinogenic potential. A population-based case–control study involving 1204 non-Asian RCC patients aged 25–74 and an equal number of sex-, age- and race-matched neighbourhood controls was conducted in Los Angeles, California, to investigate the relationship between sustained use of analgesics and risk of RCC according to major formulation categories. Detailed information on medical and medication histories, and other lifestyle factors was collected through in-person interviews. Regular use of analgesics was a significant risk factor for RCC in both men and women (odds ratio (OR) = 1.6, 95% confidence interval (CI) = 1.4–1.9 for both sexes combined). Risks were elevated across all four major classes of analgesics (aspirin, non-steroidal anti-inflammatory agents other than aspirin, acetaminophen and phenacetin). Within each class of analgesics, there was statistically significant increasing risk with increasing level of exposure. Although there was some minor variability by major class of formulation, in general individuals in the highest exposure categories exhibited approximately 2.5-fold increase in risk relative to non- or irregular users of analgesics. Subjects who took one regular-strength (i.e. 325 mg) aspirin a day or less for cardiovascular disease prevention were not at an increased risk of RCC (OR = 0.9, 95% CI = 0.6–1.4). © 1999 Cancer Research Campaig

Elicitation of Neutralizing Antibodies Directed against CD4-Induced Epitope(s) Using a CD4 Mimetic Cross-Linked to a HIV-1 Envelope Glycoprotein

The identification of HIV-1 envelope glycoprotein (Env) structures that can generate broadly neutralizing antibodies (BNAbs) is pivotal to the development of a successful vaccine against HIV-1 aimed at eliciting effective humoral immune responses. To that end, the production of novel Env structure(s) that might induce BNAbs by presentation of conserved epitopes, which are otherwise occluded, is critical. Here, we focus on a structure that stabilizes Env in a conformation representative of its primary (CD4) receptor-bound state, thereby exposing highly conserved “CD4 induced” (CD4i) epitope(s) known to be important for co-receptor binding and subsequent virus infection. A CD4-mimetic miniprotein, miniCD4 (M64U1-SH), was produced and covalently complexed to recombinant, trimeric gp140 envelope glycoprotein (gp140) using site-specific disulfide linkages. The resulting gp140-miniCD4 (gp140-S-S-M64U1) complex was recognized by CD4i antibodies and the HIV-1 co-receptor, CCR5. The gp140-miniCD4 complex elicited the highest titers of CD4i binding antibodies as well as enhanced neutralizing antibodies against Tier 1 viruses as compared to gp140 protein alone following immunization of rabbits. Neutralization against HIV-27312/V434M and additional serum mapping confirm the specific elicitation of antibodies directed to the CD4i epitope(s). These results demonstrate the utility of structure-based approach in improving immunogenic response against specific region, such as the CD4i epitope(s) here, and its potential role in vaccine application

A Directed Molecular Evolution Approach to Improved Immunogenicity of the HIV-1 Envelope Glycoprotein

A prophylactic vaccine is needed to slow the spread of HIV-1 infection. Optimization of the wild-type envelope glycoproteins to create immunogens that can elicit effective neutralizing antibodies is a high priority. Starting with ten genes encoding subtype B HIV-1 gp120 envelope glycoproteins and using in vitro homologous DNA recombination, we created chimeric gp120 variants that were screened for their ability to bind neutralizing monoclonal antibodies. Hundreds of variants were identified with novel antigenic phenotypes that exhibit considerable sequence diversity. Immunization of rabbits with these gp120 variants demonstrated that the majority can induce neutralizing antibodies to HIV-1. One novel variant, called ST-008, induced significantly improved neutralizing antibody responses when assayed against a large panel of primary HIV-1 isolates. Further study of various deletion constructs of ST-008 showed that the enhanced immunogenicity results from a combination of effective DNA priming, an enhanced V3-based response, and an improved response to the constant backbone sequences

Biofabrication: an overview of the approaches used for printing of living cells

The development of cell printing is vital for establishing biofabrication approaches as clinically relevant tools. Achieving this requires bio-inks which must not only be easily printable, but also allow controllable and reproducible printing of cells. This review outlines the general principles and current progress and compares the advantages and challenges for the most widely used biofabrication techniques for printing cells: extrusion, laser, microvalve, inkjet and tissue fragment printing. It is expected that significant advances in cell printing will result from synergistic combinations of these techniques and lead to optimised resolution, throughput and the overall complexity of printed constructs