Hyaluronic Acid: A Powerful Biomolecule with Wide-Ranging Applications-A Comprehensive Review.

Hyaluronic acid (HA) is a glycosaminoglycan widely distributed in the human body, especially in body fluids and the extracellular matrix of tissues. It plays a crucial role not only in maintaining tissue hydration but also in cellular processes such as proliferation, differentiation, and the inflammatory response. HA has demonstrated its efficacy as a powerful bioactive molecule not only for skin antiaging but also in atherosclerosis, cancer, and other pathological conditions. Due to its biocompatibility, biodegradability, non-toxicity, and non-immunogenicity, several HA-based biomedical products have been developed. There is an increasing focus on optimizing HA production processes to achieve high-quality, efficient, and cost-effective products. This review discusses HA's structure, properties, and production through microbial fermentation. Furthermore, it highlights the bioactive applications of HA in emerging sectors of biomedicine.

Drosophila melanogaster Uncoupling Protein-4A (UCP4A) Catalyzes a Unidirectional Transport of Aspartate.

Uncoupling proteins (UCPs) form a distinct subfamily of the mitochondrial carrier family (MCF) SLC25. Four UCPs, DmUCP4A-C and DmUCP5, have been identified in Drosophila melanogaster on the basis of their sequence homology with mammalian UCP4 and UCP5. In a Parkinson's disease model, DmUCP4A showed a protective role against mitochondrial dysfunction, by increasing mitochondrial membrane potential and ATP synthesis. To date, DmUCP4A is still an orphan of a biochemical function, although its possible involvement in mitochondrial uncoupling has been ruled out. Here, we show that DmUCP4A expressed in bacteria and reconstituted in phospholipid vesicles catalyzes a unidirectional transport of aspartate, which is saturable and inhibited by mercurials and other mitochondrial carrier inhibitors to various degrees. Swelling experiments carried out in yeast mitochondria have demonstrated that the unidirectional transport of aspartate catalyzed by DmUCP4 is not proton-coupled. The biochemical function of DmUCP4A has been further confirmed in a yeast cell model, in which growth has required an efflux of aspartate from mitochondria. Notably, DmUCP4A is the first UCP4 homolog from any species to be biochemically characterized. In Drosophila melanogaster, DmUCP4A could be involved in the transport of aspartate from mitochondria to the cytosol, in which it could be used for protein and nucleotide synthesis, as well as in the biosynthesis of ß-alanine and N-acetylaspartate, which play key roles in signal transmission in the central nervous system.

Physical Activity and Male Reproductive Function: A New Role for Gamete Mitochondria.

Several studies demonstrated that some types of physical exercise might affect male reproductive potential, even though the potential mechanisms involved in the modulation of sperm quality remain poorly understood. Therefore, we propose a new role for gamete mitochondria as a key hub that coordinates molecular events related to the effects induced by physical exercise.

The Effect of Physical Exercise on Cognitive Impairment in Neurodegenerative Disease: From Pathophysiology to Clinical and Rehabilitative Aspects.

Neurodegenerative diseases are a group of pathologies that cause severe disability due to motor and cognitive limitations. In particular, cognitive impairment is a growing health and socioeconomic problem which is still difficult to deal with today. As there are no pharmacologically effective treatments for cognitive deficits, scientific interest is growing regarding the possible impacts of healthy lifestyles on them. In this context, physical activity is gaining more and more evidence as a primary prevention intervention, a nonpharmacological therapy and a rehabilitation tool for improving cognitive functions in neurodegenerative diseases. In this descriptive overview we highlight the neurobiological effects of physical exercise, which is able to promote neuroplasticity and neuroprotection by acting at the cytokine and hormonal level, and the consequent positive clinical effects on patients suffering from cognitive impairment.

Cyto/Biocompatibility of Dopamine Combined with the Antioxidant Grape Seed-Derived Polyphenol Compounds in Solid Lipid Nanoparticles.

Background: The loss of nigrostriatal neurons containing dopamine (DA) together with the "mitochondrial dysfunction" in midbrain represent the two main causes related to the symptoms of Parkinson's disease (PD). Hence, the aim of this investigation is to co-administer the missing DA and the antioxidant grape seed-derived proanthocyanidins (grape seed extract, GSE) in order to increase the levels of the neurotransmitter (which is unable to cross the Blood Brain Barrier) and reducing the oxidative stress (OS) related to PD, respectively. Methods: For this purpose, we chose Solid Lipid Nanoparticles (SLN), because they have been already proven to increase DA uptake in the brain. DA-SLN adsorbing GSE (GSE/DA-SLN) were formulated and subjected to physico-chemical characterization, and their cytocompatibility and protection against OS were examined. Results: GSE was found on SLN surface and release studies evidenced the efficiency of GSE in preventing DA autoxidation. Furthermore, SLN showed high mucoadhesive strength and were found not cytotoxic to both primary Olfactory Ensheathing and neuroblastoma SH-SY5Y cells by MTT test. Co-administration of GSE/DA-SLN and the OS-inducing neurotoxin 6-hydroxydopamine (100 µM) resulted in an increase of SH-SY5Y cell viability. Conclusions: Hence, SLN formulations containing DA and GSE may constitute interesting candidates for non-invasive nose-to-brain delivery.

Drosophila melanogaster Mitochondrial Carriers: Similarities and Differences with the Human Carriers.

Mitochondrial carriers are a family of structurally related proteins responsible for the exchange of metabolites, cofactors and nucleotides between the cytoplasm and mitochondrial matrix. The in silico analysis of the Drosophila melanogaster genome has highlighted the presence of 48 genes encoding putative mitochondrial carriers, but only 20 have been functionally characterized. Despite most Drosophila mitochondrial carrier genes having human homologs and sharing with them 50% or higher sequence identity, D. melanogaster genes display peculiar differences from their human counterparts: (1) in the fruit fly, many genes encode more transcript isoforms or are duplicated, resulting in the presence of numerous subfamilies in the genome; (2) the expression of the energy-producing genes in D. melanogaster is coordinated from a motif known as Nuclear Respiratory Gene (NRG), a palindromic 8-bp sequence; (3) fruit-fly duplicated genes encoding mitochondrial carriers show a testis-biased expression pattern, probably in order to keep a duplicate copy in the genome. Here, we review the main features, biological activities and role in the metabolism of the D. melanogaster mitochondrial carriers characterized to date, highlighting similarities and differences with their human counterparts. Such knowledge is very important for obtaining an integrated view of mitochondrial function in D. melanogaster metabolism.

An SPR based immunoassay for the sensitive detection of the soluble epithelial marker E-cadherin.

Protein biomarkers are important diagnostic tools for cancer and several other diseases. To be validated in a clinical context, a biomarker should satisfy some requirements including the ability to provide reliable information on a pathological state by measuring its expression levels. In parallel, the development of an approach capable of detecting biomarkers with high sensitivity and specificity would be ideally suited for clinical applications. Here, we performed an immune-based label free assay using Surface Plasmon Resonance (SPR)-based detection of the soluble form of E-cadherin, a cell-cell contact protein that is involved in the maintaining of tissue integrity. With this approach, we obtained a specific and quantitative detection of E-cadherin from a few hundred microliters of serum of breast cancer patients by obtaining a 10-fold enhancement in the detection limit over a traditional colorimetric ELISA.

Biochemical characterization of a new mitochondrial transporter of dephosphocoenzyme A in Drosophila melanogaster.

CoA is an essential cofactor that holds a central role in cell metabolism. Although its biosynthetic pathway is conserved across the three domains of life, the subcellular localization of the eukaryotic biosynthetic enzymes and the mechanism behind the cytosolic and mitochondrial CoA pools compartmentalization are still under debate. In humans, the transport of CoA across the inner mitochondrial membrane has been ascribed to two related genes, SLC25A16 and SLC25A42 whereas in D. melanogaster genome only one gene is present, CG4241, phylogenetically closer to SLC25A42. CG4241 encodes two alternatively spliced isoforms, dPCoAC-A and dPCoAC-B. Both isoforms were expressed in Escherichia coli, but only dPCoAC-A was successfully reconstituted into liposomes, where transported dPCoA and, to a lesser extent, ADP and dADP but not CoA, which was a powerful competitive inhibitor. The expression of both isoforms in a Saccharomyces cerevisiae strain lacking the endogenous putative mitochondrial CoA carrier restored the growth on respiratory carbon sources and the mitochondrial levels of CoA. The results reported here and the proposed subcellular localization of some of the enzymes of the fruit fly CoA biosynthetic pathway, suggest that dPCoA may be synthesized and phosphorylated to CoA in the matrix, but it can also be transported by dPCoAC to the cytosol, where it may be phosphorylated to CoA by the monofunctional dPCoA kinase. Thus, dPCoAC may connect the cytosolic and mitochondrial reactions of the CoA biosynthetic pathway without allowing the two CoA pools to get in contact.

Characterization of Human and Yeast Mitochondrial Glycine Carriers with Implications for Heme Biosynthesis and Anemia.

Heme is an essential molecule in many biological processes, such as transport and storage of oxygen and electron transfer as well as a structural component of hemoproteins. Defects of heme biosynthesis in developing erythroblasts have profound medical implications, as represented by sideroblastic anemia. The synthesis of heme requires the uptake of glycine into the mitochondrial matrix where glycine is condensed with succinyl coenzyme A to yield δ-aminolevulinic acid. Herein we describe the biochemical and molecular characterization of yeast Hem25p and human SLC25A38, providing evidence that they are mitochondrial carriers for glycine. In particular, the hem25Δ mutant manifests a defect in the biosynthesis of δ-aminolevulinic acid and displays reduced levels of downstream heme and mitochondrial cytochromes. The observed defects are rescued by complementation with yeast HEM25 or human SLC25A38 genes. Our results identify new proteins in the heme biosynthetic pathway and demonstrate that Hem25p and its human orthologue SLC25A38 are the main mitochondrial glycine transporters required for heme synthesis, providing definitive evidence of their previously proposed glycine transport function. Furthermore, our work may suggest new therapeutic approaches for the treatment of congenital sideroblastic anemia.

New insights about the structural rearrangements required for substrate translocation in the bovine mitochondrial oxoglutarate carrier.

The oxoglutarate carrier (OGC) belongs to the mitochondrial carrier family and plays a key role in important metabolic pathways. Here, site-directed mutagenesis was used to conservatively replace lysine 122 by arginine, in order to investigate new structural rearrangements required for substrate translocation. K122R mutant was kinetically characterized, exhibiting a significant Vmax reduction with respect to the wild-type (WT) OGC, whereas Km value was unaffected, implying that this substitution does not interfere with 2-oxoglutarate binding site. Moreover, K122R mutant was more inhibited by several sulfhydryl reagents with respect to the WT OGC, suggesting that the reactivity of some cysteine residues towards these Cys-specific reagents is increased in this mutant. Different sulfhydryl reagents were employed in transport assays to test the effect of the cysteine modifications on single-cysteine OGC mutants named C184, C221, C224 (constructed in the WT background) and K122R/C184, K122R/C221, K122R/C224 (constructed in the K122R background). Cysteines 221 and 224 were more deeply influenced by some sulfhydryl reagents in the K122R background. Furthermore, the presence of 2-oxoglutarate significantly enhanced the degree of inhibition of K122R/C221, K122R/C224 and C224 activity by the sulfhydryl reagent 2-Aminoethyl methanethiosulfonate hydrobromide (MTSEA), suggesting that cysteines 221 and 224, together with K122, take part to structural rearrangements required for the transition from the c- to the m-state during substrate translocation. Our results are interpreted in the light of the homology model of BtOGC, built by using as a template the X-ray structure of the bovine ADP/ATP carrier isoform 1 (AAC1).

Mitochondrial glutamate carriers from Drosophila melanogaster: biochemical, evolutionary and modeling studies.

The mitochondrial carriers are members of a family of transport proteins that mediate solute transport across the inner mitochondrial membrane. Two isoforms of the glutamate carriers, GC1 and GC2 (encoded by the SLC25A22 and SLC25A18 genes, respectively), have been identified in humans. Two independent mutations in SLC25A22 are associated with severe epileptic encephalopathy. In the present study we show that two genes (CG18347 and CG12201) phylogenetically related to the human GC encoding genes are present in the D. melanogaster genome. We have functionally characterized the proteins encoded by CG18347 and CG12201, designated as DmGC1p and DmGC2p respectively, by overexpression in Escherichia coli and reconstitution into liposomes. Their transport properties demonstrate that DmGC1p and DmGC2p both catalyze the transport of glutamate across the inner mitochondrial membrane. Computational approaches have been used in order to highlight residues of DmGC1p and DmGC2p involved in substrate binding. Furthermore, gene expression analysis during development and in various adult tissues reveals that CG18347 is ubiquitously expressed in all examined D. melanogaster tissues, while the expression of CG12201 is strongly testis-biased. Finally, we identified mitochondrial glutamate carrier orthologs in 49 eukaryotic species in order to attempt the reconstruction of the evolutionary history of the glutamate carrier function. Comparison of the exon/intron structure and other key features of the analyzed orthologs suggests that eukaryotic glutamate carrier genes descend from an intron-rich ancestral gene already present in the common ancestor of lineages that diverged as early as bilateria and radiata.

Mitochondrial tricarboxylate and dicarboxylate-tricarboxylate carriers: from animals to plants.

The citrate carrier (CiC), characteristic of animals, and the dicarboxylate-tricarboxylate carrier (DTC), characteristic of plants and protozoa, belong to the mitochondrial carrier protein family whose members are responsible for the exchange of metabolites, cofactors, and nucleotides between the cytoplasm and the mitochondrial matrix. Most of the functional data on these transporters are obtained from the studies performed with the protein purified from rat, eel yeast, and maize mitochondria or recombinant proteins from different sources incorporated into phospholipid vesicles (liposomes). The functional data indicate that CiC is responsible for the efflux of acetyl-CoA from the mitochondria to the cytosol in the form of citrate, the primer for fatty acid, cholesterol synthesis, and histone acetylation. Like the CiC, the citrate exported by DTC from the mitochondria to the cytosol in exchange for oxaloacetate can be cleaved by citrate lyase to acetyl-CoA and oxaloacetate and used for fatty acid elongation and isoprenoid synthesis. In addition to its role in fatty acid synthesis, CiC is involved in other processes such as gluconeogenesis, insulin secretion, inflammation, and cancer progression, whereas DTC is involved in the production of glycerate, nitrogen assimilation, ripening of fruits, ATP synthesis, and sustaining of respiratory flux in fruit cells. This review provides an assessment of the current understanding of CiC and DTC structural and biochemical characteristics, underlying the structure-function relationship of these carriers. Furthermore, a phylogenetic relationship between CiC and DTC is proposed.

Neuroreceptor Inhibition by Clozapine Triggers Mitohormesis and Metabolic Reprogramming in Human Blood Cells.

The antipsychotic drug clozapine demonstrates superior efficacy in treatment-resistant schizophrenia, but its intracellular mode of action is not completely understood. Here, we analysed the effects of clozapine (2.5-20 µM) on metabolic fluxes, cell respiration, and intracellular ATP in human HL60 cells. Some results were confirmed in leukocytes of clozapine-treated patients. Neuroreceptor inhibition under clozapine reduced Akt activation with decreased glucose uptake, thereby inducing ER stress and the unfolded protein response (UPR). Metabolic profiling by liquid-chromatography/mass-spectrometry revealed downregulation of glycolysis and the pentose phosphate pathway, thereby saving glucose to keep the electron transport chain working. Mitochondrial respiration was dampened by upregulation of the F0F1-ATPase inhibitory factor 1 (IF1) leading to 30-40% lower oxygen consumption in HL60 cells. Blocking IF1 expression by cotreatment with epigallocatechin-3-gallate (EGCG) increased apoptosis of HL60 cells. Upregulation of the mitochondrial citrate carrier shifted excess citrate to the cytosol for use in lipogenesis and for storage as triacylglycerol in lipid droplets (LDs). Accordingly, clozapine-treated HL60 cells and leukocytes from clozapine-treated patients contain more LDs than untreated cells. Since mitochondrial disturbances are described in the pathophysiology of schizophrenia, clozapine-induced mitohormesis is an excellent way to escape energy deficits and improve cell survival.

The mitochondrial aspartate/glutamate carrier does not transport GABA.

É£-aminobutyric acid (GABA) is a four­carbon amino acid acting as the main inhibitory transmitter in the invertebrate and vertebrate nervous systems. The metabolism of GABA is well compartmentalized in the cell and the uptake of cytosolic GABA into the mitochondrial matrix is required for its degradation. A previous study carried out in the fruit fly Drosophila melanogaster indicated that the mitochondrial aspartate/glutamate carrier (AGC) is responsible for mitochondrial GABA accumulation. Here, we investigated the transport of GABA catalysed by the human and D. melanogaster AGC proteins through a well-established method for the study of the substrate specificity and the kinetic parameters of the mitochondrial carriers. In this experimental system, the D. melanogaster spliced AGC isoforms (Aralar1-PA and Aralar1-PE) and the human AGC isoforms (AGC1/aralar1 and AGC2/citrin) are unable to transport GABA both in homo- and in hetero-exchange with either glutamate or aspartate, i.e. the canonical substrates of AGC. Moreover, GABA has no inhibitory effect on the exchange activities catalysed by the investigated AGCs. Our data demonstrate that AGC does not transport GABA and the molecular identity of the GABA transporter in human and D. melanogaster mitochondria remains unknown.

Human mitochondrial uncoupling protein 3 functions as a metabolite transporter.

Since its discovery, a major debate about mitochondrial uncoupling protein 3 (UCP3) has been whether its metabolic actions result primarily from mitochondrial inner membrane proton transport, a process that decreases respiratory efficiency and ATP synthesis. However, UCP3 expression and activity are induced by conditions that would seem at odds with inefficient 'uncoupled' respiration, including fasting and exercise. Here, we demonstrate that the bacterially expressed human UCP3, reconstituted into liposomes, catalyses a strict exchange of aspartate, malate, sulphate and phosphate. The R282Q mutation abolishes the transport activity of the protein. Although the substrate specificity and inhibitor sensitivity of UCP3 display similarity with that of its close homolog UCP2, the two proteins significantly differ in their transport mode and kinetic constants.

Generation of a Yeast Cell Model Potentially Useful to Identify the Mammalian Mitochondrial N-Acetylglutamate Transporter.

The human mitochondrial carrier family (MCF) consists of 53 members. Approximately one-fifth of them are still orphans of a function. Most mitochondrial transporters have been functionally characterized by reconstituting the bacterially expressed protein into liposomes and transport assays with radiolabeled compounds. The efficacy of this experimental approach is constrained to the commercial availability of the radiolabeled substrate to be used in the transport assays. A striking example is that of N-acetylglutamate (NAG), an essential regulator of the carbamoyl synthetase I activity and the entire urea cycle. Mammals cannot modulate mitochondrial NAG synthesis but can regulate the levels of NAG in the matrix by exporting it to the cytosol, where it is degraded. The mitochondrial NAG transporter is still unknown. Here, we report the generation of a yeast cell model suitable for identifying the putative mammalian mitochondrial NAG transporter. In yeast, the arginine biosynthesis starts in the mitochondria from NAG which is converted to ornithine that, once transported into cytosol, is metabolized to arginine. The deletion of ARG8 makes yeast cells unable to grow in the absence of arginine since they cannot synthetize ornithine but can still produce NAG. To make yeast cells dependent on a mitochondrial NAG exporter, we moved most of the yeast mitochondrial biosynthetic pathway to the cytosol by expressing four E. coli enzymes, argB-E, able to convert cytosolic NAG to ornithine. Although argB-E rescued the arginine auxotrophy of arg8∆ strain very poorly, the expression of the bacterial NAG synthase (argA), which would mimic the function of a putative NAG transporter increasing the cytosolic levels of NAG, fully rescued the growth defect of arg8∆ strain in the absence of arginine, demonstrating the potential suitability of the model generated.

Clinical and Biochemical Implications of Hyaluronic Acid in Musculoskeletal Rehabilitation: A Comprehensive Review.

Hyaluronic acid (HA) naturally occurs as a biopolymer in the human body, primarily in connective tissues like joints and skin. Functioning as a vital element of synovial fluid, it lubricates joints, facilitating fluid movement and diminishing bone friction to protect articular well-being. Its distinctive attributes encompass notable viscosity and water retention capacities, ensuring flexibility and absorbing shock during motion. Furthermore, HA has gained significant attention for its potential benefits in various medical applications, including rehabilitation. Ongoing research explores its properties and functions, especially its biomedical applications in several clinical trials, with a focus on its role in improving rehabilitation outcomes. But the clinical and biochemical implications of HA in musculoskeletal rehabilitation have yet to be fully explored. This review thoroughly investigates the properties and functions of HA while highlighting its biomedical applications in different clinical trials, with a special emphasis on its role in rehabilitation. The presented findings provide evidence that HA, as a natural substance, enhances the outcomes of musculoskeletal rehabilitation through its exceptional mechanical and biochemical effects.

A novel subfamily of mitochondrial dicarboxylate carriers from Drosophila melanogaster: biochemical and computational studies.

The dicarboxylate carrier is an important member of the mitochondrial carrier family, which catalyzes an electroneutral exchange across the inner mitochondrial membrane of dicarboxylates for inorganic phosphate and certain sulfur-containing compounds. Screening of the Drosophila melanogaster genome revealed the presence of a mitochondrial carrier subfamily constituted by four potential homologs of mammalian and yeast mitochondrial dicarboxylate carriers designated as DmDic1p, DmDic2p, DmDic3p, and DmDic4p. In this paper, we report that DmDIC1 is broadly expressed at comparable levels in all development stages investigated whereas DmDIC3 and DmDIC4 are expressed only in the pupal stage, no transcripts are detectable for DmDIC2. All expressed proteins are localized in mitochondria. The transport activity of DmDic1-3-4 proteins has been investigated by reconstitution of recombinant purified protein into liposomes. DmDic1p is a typical dicarboxylate carrier showing similar substrate specificity and inhibitor sensitivity as mammalian and yeast mitochondrial dicarboxylate carriers. DmDic3p seems to be an atypical dicarboxylate carrier being able to transport only inorganic phosphate and certain sulfur-containing compounds. No transport activity was observed for DmDic4p. The biochemical results have been supported at molecular level by computing the protein structures and by structural alignments. All together these results indicate that D. melanogaster dicarboxylate carriers form a protein subfamily but the modifications in the amino acids sequences are indicative of specialized functions.

Oxidized Alginate Dopamine Conjugate: A Study to Gain Insight into Cell/Particle Interactions.

Background: We had previously synthetized a macromolecular prodrug consisting of oxidized Alginate and dopamine (AlgOx-Da) for a potential application in Parkinson disease (PD). Methods: In the present work, we aimed at gaining an insight into the interactions occurring between AlgOx-Da and SH-SY5Y neuronal cell lines in view of further studies oriented towards PD treatment. With the scope of ascertaining changes in the external and internal structure of the cells, multiple methodologies were adopted. Firstly, fluorescently labeled AlgOx-Da conjugate was synthetized in the presence of fluorescein 5(6)-isothiocyanate (FITC), providing FITC-AlgOx-Da, which did not alter SH-SY5Y cell viability according to the sulforhodamine B test. Furthermore, the uptake of FITC-AlgOx-Da by the SH-SY5Y cells was studied using scanning near-field optical microscopy and assessments of cell morphology over time were carried out using atomic force microscopy. Results: Notably, the AFM methodology confirmed that no relevant damage occurred to the neuronal cells. Regarding the effects of DA on the intracellular reactive oxygen species (ROS) production, AlgOx-Da reduced them in comparison to free DA, while AlgOx did almost not influence ROS production. Conclusions: these findings seem promising for designing in vivo studies aiming at administering Oxidized Alginate Dopamine Conjugate for PD treatment.

The Encapsulation of Citicoline within Solid Lipid Nanoparticles Enhances Its Capability to Counteract the 6-Hydroxydopamine-Induced Cytotoxicity in Human Neuroblastoma SH-SY5Y Cells.

(1) Backgrond: Considering the positive effects of citicoline (CIT) in the management of some neurodegenerative diseases, the aim of this work was to develop CIT-Loaded Solid Lipid Nanoparticles (CIT-SLNs) for enhancing the therapeutic use of CIT in parkinsonian syndrome; (2) Methods: CIT-SLNs were prepared by the melt homogenization method using the self-emulsifying lipid Gelucire® 50/13 as lipid matrix. Solid-state features on CIT-SLNs were obtained with FT-IR, thermal analysis (DSC) and X-ray powder diffraction (XRPD) studies. (3) Results: CIT-SLNs showed a mean diameter of 201 nm, -2.20 mV as zeta potential and a high percentage of entrapped CIT. DSC and XRPD analyses evidenced a greater amorphous state of CIT in CIT-SLNs. On confocal microscopy, fluorescent SLNs replacing unlabeled CIT-SLNs released the dye selectively in the cytoplasm. Biological evaluation showed that pre-treatment of SH-SY5Y dopaminergic cells with CIT-SLNs (50 µM) before the addition of 40 µM 6-hydroxydopamine (6-OHDA) to mimic Parkinson's disease's degenerative pathways counteracts the cytotoxic effects induced by the neurotoxin, increasing cell viability with the consistent maintenance of both nuclear and cell morphology. In contrast, pre-treatment with CIT 50 and 60 µM or plain SLNs for 2 h followed by 6-OHDA (40 µM) did not significantly influence cell viability. (4) Conclusions: These data suggest an enhanced protection exerted by CIT-SLNs with respect to free CIT and prompt further investigation of possible molecular mechanisms that underlie this difference.