Protein Aggregation Research Articles

Immunotherapy in Alzheimer's Disease: Current Status and Future Directions.

Alzheimer's disease (AD) is a progressive neurological disorder characterized by memory loss, cognitive decline, and behavioral changes. Immunotherapy aims to harness the immune system to target the underlying pathology of AD and has shown promise as a disease-modifying treatment for AD. By focusing on the underlying disease pathogenesis and encouraging the removal of abnormal protein aggregates in the brain, immunotherapy shows promise as a potential treatment for AD. The development of immunotherapy for AD began with early attempts to use antibodies to target beta-amyloid. The amyloid hypothesis which suggests that the accumulation of beta-amyloid in the brain triggers the pathological cascade that leads to AD has been a driving force behind the development of immunotherapy for AD. However, recent clinical trials of monoclonal antibodies targeting amyloid-β have shown mixed results, highlighting the need for further research into alternative immunotherapy approaches. Additionally, the safety and efficacy of immunotherapy for AD remain an area of active investigation. Some immunotherapeutic approaches have shown promise, while others have been associated with significant side effects, including inflammation of the brain. Sleep has a significant impact on various physiological processes, including the immune system, and has been linked to the pathogenesis of AD. Thus, improving sleep quality and duration may benefit the immune system and potentially enhance the effectiveness of immunotherapeutic approaches for AD. In this review, we discussed the promises of immunotherapy as a disease-modifying treatment for AD as well as possible methods to improve the efficacy and safety of immunotherapy to achieve better therapeutic outcomes.

Multifunctional mesoporous nanoselenium delivery of metformin breaks the vicious cycle of neuroinflammation and ROS, promotes microglia regulation and alleviates Alzheimer's disease

Clinical trials based on a single molecular target continue to fail, and the adverse effects of Aβ protein aggregation and neuroinflammation need to be solved and treatment of Alzheimer's disease. Herein, by designed a nano-sized flower mesoporous selenium transport carrier (Met@MSe@Tf) with high enzyme-like activity, metformin (Met) was loaded, and transferrin (Tf) was modified to bind to transferrin receptor to promote receptor-mediated transport across the BBB. In the AD lesion environment, with the acidic environment response dissociation, promote the release of metformin by nanoflower to achieve therapeutic effect in the brain lesion site. Metformin, a major anti-diabetic drug in diabetic metabolism, has been found to be a promising new therapeutic target in neurodegenerative diseases. Further studies showed that the metformin drug release from the designed and synthesized transport nanoparticles showed high intrinsic activity and the ability to degrade the substrate involved, especially the degradation of Aβ deposition in the cortex and hippocampus, increased the phagocytosis of microglia, thus relieving neuroinflammation simultaneously. Collectively, in vivo experiments demonstrated that Met@MSe@Tf significantly increased the number of NeuN-positive neurons in the hippocampus of AD mice, promoted neurovascular normalization in the brain, and improved cognitive dysfunction in AD transgenic AD mice. Thus, it provides a preclinical proof of concept for the construction of a highly modular accurate drug delivery platform for Alzheimer's disease.

Proteostasis and Its Role in Disease Development.

Proteostasis (protein homeostasis) refers to the general biological process that maintains the proper balance between the synthesis of proteins, their folding, trafficking, and degradation. It ensures proteins are functional, locally distributed, and appropriately folded inside cells. Genetic information enclosed in mRNA is translated into proteins. To ensure newly synthesized proteins take on the exact three-dimensional conformation, molecular chaperones assist in proper folding. Misfolded proteins can be refolded or targeted for elimination to stop aggregation. Cells utilize different degradation pathways, for instance, the ubiquitin-proteasome system, the autophagy-lysosome pathway, and the unfolded protein response, to degrade unwanted or damaged proteins. Quality control systems of the cell monitor the folding of proteins. These checkpoint mechanisms are aimed at degrading or refolding misfolded or damaged proteins. Under stress response pathways, such as heat shock response and unfolded protein response, which are triggered under conditions that perturb proteostasis, the capacity for folding is increased, and degradation pathways are activated to help cells handle stressful conditions. The deregulation of proteostasis is implicated in a variety of illnesses, comprising cancer, metabolic diseases, cardiovascular diseases, and neurological disorders. Therapeutic strategies with a deeper insight into the mechanism of proteostasis are crucial for the treatment of illnesses linked with proteostasis and to support cellular health. Thus, proteostasis is required not only for the maintenance of cellular homeostasis and function but also for proper protein function and prevention of injurious protein aggregation. In this review, we have covered the concept of proteostasis, its mechanism, and how disruptions to it can result in a number of disorders.

Gelation properties and swallowing characteristics of heat-induced whey protein isolate/chia seed gum composite gels as dysphagia food

Soft gels based on protein-polysaccharide composite systems play a crucial role in the dietary management of people with dysphagia. The effect of chia seed gum (CSG) on the gelling and swallowing properties of heat-induced whey protein isolate (WPI) gels (3.125–75 mg/mL) was investigated. The results showed that adding CSG reduced the gelation concentration of WPI and weak gels could form at 12.5 mg/mL WPI concentration. In addition, the viscoelasticity and water-binding capacity of the WPI/CSG composite gels were gradually enhanced with increasing WPI concentrations. The WPI/CSG composite systems can be classified as level 2–5 dysphagia-oriented foods according to the International Dysphagia Diet Standardization Initiative (IDDSI) framework. The incorporation of CSG promoted the cross-linking of protein aggregates and the formation of compact and continuous network structures, resulting in improved gelling properties of composite systems. This study contributes to the development of novel soft gel-type dysphagia foods with better textural characteristics.

A Randomized-Controlled Trial Targeting Cognition in Early Alzheimer's Disease by Improving Sleep with Trazodone (REST).

Alzheimer's disease (AD) is a leading cause of mortality and morbidity among aging populations worldwide. Despite arduous research efforts, treatment options for this devastating neurodegenerative disease are limited. Sleep disturbances, through their link to changes in neural excitability and impaired clearance of interstitial abnormal protein aggregates, are a key risk factor for the development of AD. Research also suggests that the neuroprotective effects of sleep are particularly active during slow wave sleep. Given the strong link between sleep disturbance and AD, targeting sleep in the prodromal stages of AD, such as in mild cognitive impairment (MCI), represents a promising avenue for slowing the onset of AD-related cognitive decline. In efforts to improve sleep in older individuals, several pharmacologic approaches have been employed, but many pose safety risks, concern for worsening cognitive function, and fail to effectively target slow wave sleep. Trazodone, a safe and widely used drug in the older adult population, has shown promise in inducing slow wave sleep in older adults, but requires more rigorous research to understand its effects on sleep and cognition in the prodromal stages of AD. In this review, we present the rationale and study design for our randomized, double-bind, placebo-controlled, crossover trial (NCT05282550) investigating the effects of trazodone on sleep and cognition in 100 older adults with amnestic MCI and sleep complaints.

The neurotrophic factor MANF regulates autophagy and lysosome function to promote proteostasis in Caenorhabditis elegans

The conserved mesencephalic astrocyte-derived neurotrophic factor (MANF) is known for protecting dopaminergic neurons and functioning in various other tissues. Previously, we showed that Caenorhabditis elegans manf-1 null mutants exhibit defects such as increased endoplasmic reticulum (ER) stress, dopaminergic neurodegeneration, and abnormal protein aggregation. These findings suggest an essential role for MANF in cellular processes. However, the mechanisms by which intracellular and extracellular MANF regulate broader cellular functions remain unclear. We report a unique mechanism of action for MANF-1 that involves the transcription factor HLH-30/TFEB-mediated signaling to regulate autophagy and lysosomal function. Multiple transgenic strains overexpressing MANF-1 showed extended lifespan of animals, reduced protein aggregation, and improved neuronal survival. Using fluorescently tagged MANF-1, we observed tissue-specific localization of the protein, which was dependent on the ER retention signal. Further subcellular analysis showed that MANF-1 localizes within cells to the lysosomes and utilizes the endosomal pathway. Consistent with the lysosomal localization, our transcriptomic study of MANF-1 and analyses of autophagy regulators demonstrated that MANF-1 promotes proteostasis by regulating autophagic flux and lysosomal activity. Collectively, our findings establish MANF as a critical regulator of stress response, proteostasis, and aging.

Analyzing Amylin Aggregation Inhibition Through Quantum Dot Fluorescence Imaging.

Protein aggregation is associated with various diseases caused by protein misfolding. Among them, amylin deposition is a prominent feature of type 2 diabetes. At present, the mechanism of amylin aggregation remains unclear, and this has hindered the treatment of type 2 diabetes. In this study, we analyzed the aggregation process of amylin using the quantum dot (QD) imaging method. QD fluorescence imaging revealed that in the presence of 100 μM amylin, aggregates appeared after 12 h of incubation, while a large number of aggregates formed after 24 h of incubation, with a standard deviation (SD) value of 5.435. In contrast, 50 μM amylin did not induce the formation of aggregates after 12 h of incubation, although a large number of aggregates were observed after 24 h of incubation, with an SD value of 2.883. Confocal laser microscopy observations revealed that these aggregates were deposited in three dimensions. Transmission electron microscopy revealed that amylin existed as misfolded fibrils in vitro and that QDs were uniformly bound to the amylin fibrils. In addition, using a microliter-scale high-throughput screening (MSHTS) system, we found that rosmarinic acid, a polyphenol, inhibited amylin aggregation at a half-maximal effective concentration of 852.8 μM. These results demonstrate that the MSHTS system is a powerful tool for evaluating the inhibitory activity of amylin aggregation. Our findings will contribute to the understanding of the pathogenesis of amylin-related diseases and the discovery of compounds that may be useful in the treatment and prevention of these diseases.

Properties of Rennet Gels from Retentate Produced by Cold Microfiltration of Heat-Treated and Microfiltered Skim Milk.

This study investigated the production of rennet gels from β-casein-depleted retentates obtained through cold microfiltration (MF) of skim milk (SM) that was treated beforehand to ensure microbial safety. The treatments included thermization (65 °C, 20 s), pasteurization (72 °C, 15 s), and microfiltration (50 °C; 1.4 μm pore size). The reduction in β-casein content was 0.98, 0.51 and 0.90%, respectively. All treatments resulted in the partial aggregation of serum proteins, which were slightly concentrated in the retentates obtained post cold MF process. This aggregation, along with concentration effect, likely inhibited β-casein dissociation from casein micelles and permeation, particularly in pasteurized milk. Renneting and coagulation properties of the retentates were comparable to those of the respective SM samples, with no significant differences in syneresis, water-holding capacity, or protein hydration. Notably, the retentate from thermized SM, which showed the best performance with the highest β-casein reduction (0.98%), demonstrated shorter coagulation time compared to retentate from pasteurized milk or the corresponding unfiltered SM. Textural analysis revealed greater firmness, cohesiveness, and viscosity of retentate-based rennet gels compared to gels made from unfiltered SM, attributed to protein concentration during cold MF. Overall, this study successfully produced rennet gels from cold MF retentates without compromising their physicochemical properties.

Retraction: Short polyethylene glycol chains densely bound to soft nanotube channels for inhibition of protein aggregation.

[This retracts the article DOI: 10.1039/C6RA06793J.].

The Emerging Role of Phosphodiesterase 5 Inhibition in Neurological Disorders: The State of the Art.

Growing evidence suggests that neuroinflammation is not just a consequence of neurodegeneration in pathologies such as Alzheimer's disease, Parkinson's disease, Huntington's disease or Amyotrophic lateral sclerosis, but it is rather a determinant factor, which plays a pivotal role in the onset and progression of these disorders. Neuroinflammation can affect cells and processes in the central nervous system (CNS) as well as immune cells, and might precede protein aggregation, which is a hallmark of the neurodegenerative process. Standard treatment methods are far from being able to counteract inflammation and delay neurodegeneration. Remarkably, phosphodiesterase 5 inhibitors (PDE5is), which represent potent vasoactive drugs used as a first-line treatment for erectile dysfunction (ED), display important anti-inflammatory effects through cyclic guanosine monophosphate (cGMP) level stabilization. Since PDE5 hydrolyzes cGMP, several studies positioned PDE5 as a therapeutic target, and more specifically, PDE5is as potential alternative strategies for the treatment of a variety of neurological disorders. Indeed, PDE5is can limit neuroinflammation and enhance synaptic plasticity, with beneficial effects on cognitive function and memory. The aim of this review is to provide an overview of some of the main processes underlying neuroinflammation and neurodegeneration which may be potential targets for PDE5is, focusing on sildenafil, the most extensively studied. Current strategies using PDEis for the treatment of neurodegenerative diseases will be summarized.

Aβ-targeting synNotch Receptor for Alzheimer's Disease: Expanding Applications to Extracellular Protein Aggregates.

The synthetic Notch receptor (synNotch) system is a versatile platform that induces gene transcription in response to extracellular signals. However, its application has been largely confined to membrane-bound targets due to specific activation requirements. Whether synNotch can also target extracellular protein aggregates, such as amyloid beta (Aβ) in Alzheimer's disease (AD), is unclear. To address this, we engineered an Aβ-targeting synNotch receptor controlling the production of chimeric human-mouse versions of Lecanemab (Leqembi®) or Aducanumab (Aduhelm®), both FDA-approved antibodies for AD. We demonstrate that NIH 3T3 cells expressing this synNotch system detect and respond to extracellular Aβ aggregates by synthesizing and secreting Aducanumab or Lecanemab. These findings broaden the potential applications of synNotch, extending its targets beyond membrane-bound proteins to extracellular protein aggregates, providing obvious benefits to research in this scientific arena.

Enhanced Photodynamic Therapy for Neurodegenerative Diseases: Development of Azobenzene-Spiropyran@Gold Nanoparticles for Controlled Singlet Oxygen Generation.

The development of durable photosensitizers is pivotal for advancing phototherapeutic applications in biomedicine. Here, we introduce a core-shell azobenzene-spiropyran structure on gold nanoparticles, engineered to enhance singlet oxygen generation. These nano-photosensitizers exhibit increased structural stability and thermal resistance, as demonstrated by slowed O-N-C bond recombination dynamics via in-situ Raman spectroscopy. Notably, the in-situ formation of merocyanine and a light-induced compact shell arrangement extend its half-life from 47 minutes to over 154 hours, significantly boosting singlet oxygen output. The nano-photosensitizer also shows high biocompatibility and notably inhibits tau protein aggregation in neural cells, even with phosphatase inhibitors. Further, it promotes dendritic growth in neuro cells, doubling typical lengths. This work not only advances chemical nanotechnology but also sets a foundation for developing long-lasting phototherapy agents for treating neurodegenerative diseases.

Disruption of PHF6 Peptide Aggregation from Tau Protein: Mechanisms of Palmatine Chloride in Preventing Early PHF6 Aggregation.

The formation of neurofibrillary tangles (NFTs), composed of tau protein aggregates, is a hallmark of neurodegenerative diseases known as tauopathies, including Alzheimer's disease (AD). NFTs consist of paired helical filaments (PHFs) of tau protein with a dominant β-sheet secondary structure. Within these PHFs, the PHF6 hexapeptide (Val306-Gln-Ile-Val-Tyr-Lys311) has been commonly highlighted as a key site for tau protein nucleation. Palmatine chloride (PC) has been identified as an inhibitor of PHF6 aggregation, capable of reducing aggregation propensity at submicromolar concentrations. In pursuit of novel anti-AD drugs targeting early tau aggregation stages, we conducted an in silico study to elucidate PC's mechanism of action during PHF6 aggregation. Our observations suggest that while PHF6 can still initiate self-aggregation in the presence of PC, PC molecules subtly influence PHF6 aggregation dynamics, favoring smaller aggregates over larger complexes. The study underlined the key roles of aromatic rings in PC binding to different PHF6 aggregates by interacting through π-π stacking with the PHF6 Tyr310 side chain. The presence of aromatic rings in compounds to be able to inhibit the earlier complexation phase seems to be essential. These in silico findings lay a foundation for the design of compounds that could intervene in resolving the neurotoxicity of protein aggregates in AD.

Early steps of protein disaggregation by Hsp70 chaperone and class B J-domain proteins are shaped by Hsp110

Hsp70 is a key cellular system counteracting protein misfolding and aggregation, associated with stress, ageing, and disease. Hsp70 solubilises aggregates and aids protein refolding through substrate binding and release cycles regulated by co-chaperones: J-domain proteins (JDPs) and nucleotide exchange factors (NEFs). Here, we elucidate the collaborative impact of Hsp110 NEFs and different JDP classes throughout Hsp70-dependent aggregate processing. We show that Hsp110 plays a major role at initial stages of disaggregation, determining its final efficacy. The NEF catalyses the recruitment of thick Hsp70 assemblies onto aggregate surface, which modifies aggregates into smaller species more readily processed by chaperones. Hsp70 stimulation by Hsp110 is much stronger with class B than class A JDPs and requires the auxiliary interaction between class B JDP and the Hsp70 EEVD motif. Furthermore, we demonstrate for the first time that Hsp110 disrupts the JDP-Hsp70 interaction. Such destabilisation of chaperone complexes at the aggregate surface might improve disaggregation, but also lead to the inhibition above the sub-stoichiometric Hsp110 optimum. Thus, balanced interplay between the co-chaperones and Hsp70 is critical to unlock its disaggregating potential.

Early steps of protein disaggregation by Hsp70 chaperone and class B J-domain proteins are shaped by Hsp110.

Hsp70 is a key cellular system counteracting protein misfolding and aggregation, associated with stress, ageing, and disease. Hsp70 solubilises aggregates and aids protein refolding through substrate binding and release cycles regulated by co-chaperones: J-domain proteins (JDPs) and nucleotide exchange factors (NEFs). Here, we elucidate the collaborative impact of Hsp110 NEFs and different JDP classes throughout Hsp70-dependent aggregate processing. We show that Hsp110 plays a major role at initial stages of disaggregation, determining its final efficacy. The NEF catalyses the recruitment of thick Hsp70 assemblies onto aggregate surface, which modifies aggregates into smaller species more readily processed by chaperones. Hsp70 stimulation by Hsp110 is much stronger with class B than class A JDPs and requires the auxiliary interaction between class B JDP and the Hsp70 EEVD motif. Furthermore, we demonstrate for the first time that Hsp110 disrupts the JDP-Hsp70 interaction. Such destabilisation of chaperone complexes at the aggregate surface might improve disaggregation, but also lead to the inhibition above the sub-stoichiometric Hsp110 optimum. Thus, balanced interplay between the co-chaperones and Hsp70 is critical to unlock its disaggregating potential.

Discovery of novel substituted (Z)-N′-hydroxy-3-(3-phenylureido)benzimidamide derivatives as multifunctional molecules targeting pathological hallmarks of Alzheimer's disease

Alzheimer's disease (AD) is a neurodegenerative disorder marked by significant loss of central cholinergic neurons. This progressive deterioration leads to cognitive dysfunction and impaired motor activity, culminating in the brain cell's death at the later stages of the disease. The approved drugs for AD are limited to providing symptomatic relief for an initial period due to the multifaceted etiology of the disease. Several studies have demonstrated that rivastigmine (RIV) is a selectively potent inhibitor of butyrylcholinesterase and devoid of antioxidant, Aβ, and tau protein aggregation inhibition and anti-inflammatory properties. Therefore, to address these issues associated with RIV, novel rivastigmine-based molecules were rationally designed, synthesized, and evaluated in various in-vitro and in-vivo AD models. In in-vitro acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) inhibition studies revealed that 3q &6e as promising leads (AChE, IC50 1.72 ± 0.15, 0.91 ± 0.016 μM, BChE, IC50 6.69 ± 0.28 μM, 1.19 ± 0.026 μM, for 3q &6e, respectively). The computational studies (molecular docking and dynamics) further corroborated the in-vitro studies. Further, 3q and 6e were found to be potent antioxidants in the DPPH assay (IC50 16.15 ± 1.05 & 15.17 ± 0.07 μM, for 3q &6e, respectively). Interestingly, 3q, and 6e could effectively inhibit self-induced full-length tau and Aβ1-42 aggregation. Treatment with 3q &6e inhibited microglial activation by attenuating ROS release and mitochondrial damage. Further, 3q &6e also suppressed NLRP3 inflammasome and NF-κB expression levels in microglial cells and halted the release of pro-inflammatory cytokines in human microglial cells. Finally, 3q &6e were found to be efficacious in reversing the scopolamine-induced memory impairment in the Morris water maze test. The expression of various neuroprotection markers, such as BDNF and TRKB, was significantly overexpressed compared to the disease control group.

Structure-based computer-aided drug design to identify potential lead molecules for Asparaginyl Endopeptidase inhibitors.

The enzyme Asparaginyl Endopeptidase (AEP) is associated with proteinopathy-related pathologies such as Alzheimer's disease (AD) and Frontal Temporal Dementia (FTD). The onset of pathologies by AEP is due to cleaved fragments forming protein aggregates resulting in neurodegeneration. Unfortunately, there are no clinically approved small molecule inhibitors for AEP, and therefore, it serves as an unmet medical need for the design and development of potential novel small molecules. In developing potential inhibitors for proteolytic activity, a structured approach utilizing structure-based computer-aided drug design (SB-CADD) parameters was employed. This involved virtual high throughput screening (vHTS) across various CNS-focused databases enriched with diverse functionality. We identified the top sixty ligands based on the glide XP-docking score out of 10 million ligands. The free binding energy was then calculated using MM-GBSA for all top selected molecules which resulted in discovering that AEPI-1 to AEPI-6 (Asparaginyl Endopeptidase inhibitors) displayed high affinity towards the catalytic triad. Further investigation determined that all top six hits form stable complexes during 50 ns molecular dynamic simulations. We also observed that AEPI-2 demonstrated the highest stability within the binding pockets. Post-MD analyses such as DCCM, PCA, PDF, and ADMET properties were also evaluated. By bridging all the observations, we observed these six molecules occupy the active site of the β-helix (β1, β3, and β4) of the S1 pocket and additional binding sites in α1 and β5, suggesting its suitability as a potential candidate for drug discovery against Asparaginyl Endopeptidase.

Homologous Peptide Foldamer Promotes FUS Aggregation and Triggers Cancer Cell Death.

Fused in sarcoma (FUS), a multifunctional deoxyribonucleic acid (DNA)/ribonucleic acid (RNA)-binding protein, has been implicated in various cancer types, including sarcoma and leukemia. Despite its association with these diseases, there has been limited exploration of FUS as a cancer therapy target, primarily because its dynamic nature makes it difficult to target specifically. In this study, we explored a kind of β-sheet peptide foldamer, named β4-TAT, to influence FUS aggregation by targeting its RNA recognition motifs (RRM). This approach leverages the noncovalent interaction characteristics of peptide self-assembly processes. The β4 sequence, derived from the FUS RRM β-sheet, in combination with TAT, a peptide known for its nuclear targeting capability, enables β4-TAT to bind specifically to the analogous β4 sequence within FUS. Notably, β4-TAT effectively induces FUS aggregation within cells, leading to the death of cancer cells. Our work developed a novel peptide foldamer-based strategy for inducing protein aggregation, paving the way for innovative therapeutic approaches in targeting FUS-associated cancers.

Proteome Aggregation in Cells Derived from Amyotrophic Lateral Sclerosis Patients for Personalized Drug Evaluation.

Amyotrophic lateral sclerosis (ALS) is a progressive neurodegenerative disorder that currently lacks effective therapy. Given the heterogeneity of clinical and molecular profiles of ALS patients, personalized diagnostics and pathological characterization represent a powerful strategy to optimize patient stratification, thereby enabling personalized treatment. Immortalized lymphocytes from sporadic and genetic ALS patients recapitulate some pathological hallmarks of the disease, facilitating the fundamental task of drug screening. However, the molecular aggregation of ALS has not been characterized in this patient-derived cellular model. Indeed, protein aggregation is one of the most prominent features of neurodegenerative diseases, and therefore, models to test drugs against personalized pathological aggregation could help discover improved therapies. With this work, we aimed to characterize the aggregation profile of ALS immortalized lymphocytes and test several drug candidates with different mechanisms of action. In addition, we have evaluated the molecular aggregation in motor neurons derived from two hiPSC cell lines corresponding to ALS patients with different mutations in TARDBP. The results provide valuable insight into the different characterization of sporadic and genetic ALS patients' immortalized lymphocytes, their differential response to drug treatment, and the usefulness of proteome homeostasis characterization in patients' cells.

Exploring the interplay between zinc‐induced protein dyshomeostasis and mitochondrial dysfunction using viscosity‐sensitive sensor

AbstractMitochondria are crucial sites for protein quality control within cells. When mitochondrial stress is triggered by protein misfolding, it can accelerate abnormal protein aggregation, potentially inducing various diseases. This study developed a cascade‐responsive sensor, named AggHX, to monitor the microenvironment of protein aggregation induced by zinc (II) ions and the accompanying mitochondrial dysfunction. The AggHX consists of two key components: (1) A Zn2+ recognition group for triggering a fluorescent enhance response, and (2) a near‐infrared BODIPY scaffold that detects viscosity changes in cell aggregation via HaloTag. This sensor's mechanism of action is elucidated through photochemical and biochemical characterizations. To further investigate the relationship between protein aggregation and mitochondrial homeostasis, we employ fluorescence lifetime imaging microscopy to assess viscosity changes in protein aggregates under intracellular Zn2+ stress. This research provides insights into the dynamic behavior and spatial distribution of protein aggregates and mitochondria, contributing to a deeper understanding of their physiological roles in cellular processes and potential implications in disease pathology.