Amphipathic peptides composed of alternating hydrophobic and hydrophilic amino acids self-assemble into amyloid-inspired, β-sheet nanoribbon fibrils. Herein, we report a new fibril type that is ...formed from equimolar mixtures of enantiomeric amphipathic peptides (l- and d-(FKFE)2). Spectroscopic analysis indicates that these peptides do not self-sort and assemble into enantiomeric fibrils composed of all-l and all-d peptides, but rather coassemble into fibrils that contain alternating l- and d-peptides in a “rippled β-sheet” orientation. Isothermal titration calorimetry indicates an enthalpic advantage for rippled β-sheet coassembly compared to self-sorted β-sheet assembly of enantiomeric peptides.
Self-assembled peptide-based hydrogels are emerging materials that have been exploited for wound healing, drug delivery, tissue engineering, and other applications. In comparison to synthetic polymer ...hydrogels, supramolecular peptide-based gels have advantages in biocompatibility, biodegradability, and ease of synthesis and modification. Modification of the emergent viscoelasticity of peptide hydrogels in a stimulus responsive fashion is a longstanding goal in the development of next-generation materials. In an effort to selectively modulate hydrogel viscoelasticity, we report herein a method to enhance the elasticity of β-sheet peptide hydrogels using specific molecular recognition events between functionalized hydrogel fibrils and biomolecules. Two distinct biomolecular recognition strategies are demonstrated: oligonucleotide Watson–Crick duplex formation between peptide nucleic acid (PNA) modified fibrils with a bridging oligonucleotide and protein–ligand recognition between mannose modified fibrils with concanavalin A. These methods to modulate hydrogel elasticity should be broadly adaptable in the context of these materials to a wide variety of molecular recognition partners.
Semen-derived enhancer of viral infection (SEVI), an amyloid fibril formed from a cationic peptide fragment of prostatic acidic phosphatase (PAP), dramatically enhances the infectivity of human ...immunodeficiency virus type 1 (HIV-1). Insoluble, sedimentable fibrils contribute to SEVI-mediated enhancement of virus infection. However, the SEVI-forming PAP(248–286) peptide is able to produce infection-enhancing structures much more quickly than it forms amyloid fibrils. This suggests that soluble supramolecular assemblies may enhance HIV-1 infection. To address this question, non-SEVI amyloid-like fibrils were derived from general amphipathic peptides of sequence Ac-K
n(XKXE)
2-NH
2. These cationic peptides efficiently self-assembled to form soluble, fibril-like structures that were, in some cases, able to enhance HIV-1 infection even more efficiently than SEVI. Experiments were also performed to determine whether agents that efficiently shield the charged surface of SEVI fibrils block SEVI-mediated infection-enhancement. To do this, we generated self-assembling anionic peptides of sequence Ac-E
n(XKXE)
2-NH
2. One of these peptides completely abrogated SEVI-mediated enhancement of HIV-1 infection, without altering HIV-1 infectivity in the absence of SEVI. Collectively, these data suggest that soluble SEVI assemblies may mediate infection-enhancement, and that anionic peptide supramolecular assemblies have the potential to act as anti-SEVI microbicides.
The display of functional proteins on self-assembled peptide nanofibrils is challenging since the steric bulk of proteins attached to simple self-assembling peptides often impedes incorporation into ...nanofibrils. Herein is described a split-protein strategy to tether functional proteins to preassembled peptide nanofibrils. In this strategy, a short affinity motif peptide derived from a split protein system is appended to a self-assembly motif (the amphipathic Ac-(FKFE)
-NH
peptide) to form an affinity-assembly fusion peptide. The small size of the affinity motif allows the affinity-assembly fusion peptide to be readily incorporated into peptide nanofibrils that display the affinity motif when the affinity-assembly peptide is coassembled with Ac-(FKFE)
-NH
. Introduction of the split-protein that is complementary to the affinity motif to the assembled nanofibrils results in efficient, multivalent attachment of functional proteins to the peptide nanofibrils. This strategy is demonstrated with two split-protein systems, ribonuclease S' (RNase S') and split green fluorescent protein (GFP).
Amyloid fibrils contained in semen, known as SEVI, or semen-derived enhancer of viral infection, have been shown to increase the infectivity of HIV dramatically. However, previous work with these ...fibrils has suggested that extensive time and nonphysiologic levels of agitation are necessary to induce amyloid formation from the precursor peptide (a proteolytic cleavage product of prostatic acid phosphatase, PAP248–286). Here, we show that fibril formation by PAP248–286 is accelerated dramatically in the presence of seminal plasma (SP) and that agitation is not required for fibrillization in this setting. Analysis of the effects of specific SP components on fibril formation by PAP248–286 revealed that this effect is primarily due to the anionic buffer components of SP (notably inorganic phosphate and sodium bicarbonate). Divalent cations present in SP had little effect on the kinetics of fibril formation, but physiologic levels of Zn2+ strongly protected SEVI fibrils from degradation by seminal proteases. Taken together, these data suggest that in the in vivo environment, PAP248–286 is likely to form fibrils efficiently, thus providing an explanation for the presence of SEVI in human semen.
Background: SEVI is an amyloid fibril that enhances HIV infectivity. To date, it has been produced from its precursor peptide only under nonphysiologic conditions.
Results: Seminal plasma (SP) accelerates SEVI formation and protects SEVI from proteolytic degradation.
Conclusion: SEVI forms spontaneously from its precursor peptide under physiologic conditions in SP.
Significance: These findings may explain the presence of SEVI in human semen.
Cationic amyloid fibrils, including the Semen Enhancer of Virus Infection (SEVI), have recently been described in human semen. Simple methods for quantitating these fibrils are needed to improve our ...understanding of their biological function. We performed high-throughput screening to identify molecules that bind SEVI, and identified a small molecule (8E2), that fluoresced brightly in the presence of SEVI and other cationic fibrils. 8E2 bound SEVI with almost 40-fold greater affinity than thioflavin-T, and could efficiently detect high molecular weight fibrils in human seminal fluid.
Abstract
Currently, the incorrect judgment of burn depth remains common even among experienced surgeons. Contributing to this problem are change in burn appearance throughout the first week requiring ...periodic evaluation until a confident diagnosis can be made. To overcome these issues, we investigated the feasibility of an artificial intelligence algorithm trained with multispectral images of burn injuries to predict burn depth rapidly and accurately, including burns of indeterminate depth. In a feasibility study, 406 multispectral images of burns were collected within 72 hours of injury and then serially for up to 7 days. Simultaneously, the subject’s clinician indicated whether the burn was of indeterminate depth. The final depth of burned regions within images were agreed upon by a panel of burn practitioners using biopsies and 21-day healing assessments as reference standards. We compared three convolutional neural network architectures and an ensemble in their capability to automatically highlight areas of nonhealing burn regions within images. The top algorithm was the ensemble with 81% sensitivity, 100% specificity, and 97% positive predictive value (PPV). Its sensitivity and PPV were found to increase in a sigmoid shape during the first week postburn, with the inflection point at day 2.5. Additionally, when burns were labeled as indeterminate, the algorithm’s sensitivity, specificity, PPV, and negative predictive value were: 70%, 100%, 97%, and 100%. These results suggest multispectral imaging combined with artificial intelligence is feasible for detecting nonhealing burn tissue and could play an important role in aiding the earlier diagnosis of indeterminate burns.
Mitochondria are central organelles in cellular metabolism. Their structure is highly dynamic, allowing them to adapt to different energy requirements, to be partitioned during cell division, and to ...maintain functionality. Mitochondrial dynamics, including membrane fusion and fission reactions, are well studied in yeast and mammals but it is not known if these processes are conserved throughout eukaryotic evolution. Kinetoplastid parasites are some of the earliest-diverging eukaryotes to retain a mitochondrion. Each cell has only a single mitochondrial organelle, making them an interesting model for the role of dynamics in controlling mitochondrial architecture. We have investigated the mitochondrial division cycle in the kinetoplastid Crithidia fasciculata. The majority of mitochondrial biogenesis occurs during the G1 phase of the cell cycle, and the mitochondrion is divided symmetrically in a process coincident with cytokinesis. Live cell imaging revealed that the mitochondrion is highly dynamic, with frequent changes in the topology of the branched network. These remodeling reactions include tubule fission, fusion, and sliding, as well as new tubule formation. We hypothesize that the function of this dynamic remodeling is to homogenize mitochondrial contents and to facilitate rapid transport of mitochondria-encoded gene products from the area containing the mitochondrial nucleoid to other parts of the organelle.
Background
Although left ventricular assist device (LVAD) implantation is associated with improved heart failure survival, the impact of pre‐implantation Impella support on outcomes is unknown. We ...undertook this study to evaluate the impact of preoperative Impella support on LVAD outcomes.
Methods
We conducted a retrospective review of all Heartmate 3 LVAD implants. Primary stratification was by the need for preoperative Impella support with the 5.0/5.5 device. Longitudinal survival was assessed by the Kaplan–Meier method. Multivariable Cox proportional hazards regression models were developed to evaluate mortality. Secondary outcomes included changes in laboratory values during Impella support.
Results
From 2017 to 2021, 87 patients underwent LVAD implantation. Sixteen were supported with a single inotrope, 36 with dual inotropes, 27 with Impella, and 3 with extracorporeal membrane oxygenation (ECMO). When stratified by the need for Impella, there was no difference in survival at 30‐days (98.3 88.2–99.8% vs. 96.3 76.5–99.5%, p = .59), 1‐year (91.0 79.8–96.2 vs. 74.9 51.7–88.2, p = .10), or at 2 years (87.9 74.3–94.5 vs. 74.9 51.7–88.2, p = .15). On multivariable modeling, the need for preoperative Impella was not associated with an increased hazard of 1‐year (1.24 0.23–6.73, p = .81) or 2‐year mortality (1.05 0.21–5.19, p = .95). After 7 (5–10) days of Impella support, recipient creatinine (p < .01), creatinine clearance (p = .02), and total bilirubin (p = .053) improved and lactic acidosis resolved (p < .01).
Conclusions
Preoperative Impella support is not associated with increased short or long‐term mortality but is associated with improved renal and hepatic function as well as total body perfusion before LVAD implantation.
We used CDK4/hTERT-immortalized normal human bronchial epithelial cells (HBEC) from several individuals to study lung cancer pathogenesis by introducing combinations of common lung cancer oncogenic ...changes (p53, KRAS, and MYC) and followed the stepwise transformation of HBECs to full malignancy. This model showed that: (i) the combination of five genetic alterations (CDK4, hTERT, sh-p53, KRAS(V12), and c-MYC) is sufficient for full tumorigenic conversion of HBECs; (ii) genetically identical clones of transformed HBECs exhibit pronounced differences in tumor growth, histology, and differentiation; (iii) HBECs from different individuals vary in their sensitivity to transformation by these oncogenic manipulations; (iv) high levels of KRAS(V12) are required for full malignant transformation of HBECs, however, prior loss of p53 function is required to prevent oncogene-induced senescence; (v) overexpression of c-MYC greatly enhances malignancy but only in the context of sh-p53+KRAS(V12); (vi) growth of parental HBECs in serum-containing medium induces differentiation, whereas growth of oncogenically manipulated HBECs in serum increases in vivo tumorigenicity, decreases tumor latency, produces more undifferentiated tumors, and induces epithelial-to-mesenchymal transition (EMT); (vii) oncogenic transformation of HBECs leads to increased sensitivity to standard chemotherapy doublets; (viii) an mRNA signature derived by comparing tumorigenic versus nontumorigenic clones was predictive of outcome in patients with lung cancer. Collectively, our findings show that this HBEC model system can be used to study the effect of oncogenic mutations, their expression levels, and serum-derived environmental effects in malignant transformation, while also providing clinically translatable applications such as development of prognostic signatures and drug response phenotypes.
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