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Mono- and disubstituted 2,6-dibromo-dimethylaminophenylbuta-1,3-dienylBODIPY dyes were successfully prepared, and their in vitro photodynamic activities against MCF-7 breast cancer cells were evaluated with a Thorlabs M660L4 660 nm LED (336 J · cm[Formula: see text]. The IC[Formula: see text] value of the monophenylbuta-1,3-dienylBODIPY was ca. 2.1 [Formula: see text]M, while that of the diphenylbuta-1,3-dienylBODIPY was > 50 [Formula: see text]M. Both dyes exhibited minimal dark toxicity. The results demonstrate that monosubstituted 2,6-dibromo-dimethylaminophenylbuta-1,3-dienylBODIPY dyes merit further in-depth study for use as photosensitizer dyes in photodynamic therapy.

Tin(IV) complexes of a 4-methylthiophenyl functionalized porphyrin (1-Sn) and its corrole analogue (2-Sn) were synthesized so that their photophysicochemical properties and photodynamic activities against MCF-7 breast cancer cells could be compared. Singlet oxygen luminescence studies revealed that 1-Sn and 2-Sn have comparable [Formula: see text] values in DMF of 0.59 and 0.60, respectively, while the IC[Formula: see text] values after irradiation of MCF-7 cells for 30 min with a Thorlabs 625 nm LED (432 J · cm[Formula: see text] were determined to be 12.4 and 8.9 [Formula: see text]M. The results demonstrate that the cellular uptake of 2-Sn and its molar absorptivity at the irradiation wavelength play a crucial role during in vitro cytotoxicity studies.

The hypoxic tumour microenvironment and poor spatiotemporal localization of photosensitizers are two significant obstacles that limit practical applications of photodynamic therapy. In response, a biocompatible, light-activatable liposome integrated with both a zinc phthalocyanine photodynamic component and Pt nanoparticles-decorated with MnO2 catalase-mimicking component are engineered. This multifunctional system was rationally designed using unsaturated phospholipids to achieve on-demand drug release following light irradiation. Specificity was achieved by folic acid functionalization resulting in folate-modified liposomes (FTLiposomes). We demonstrated its specific uptake by fluorescence imaging using folate receptor (FR) overexpressing HeLa and MCF-7 cells as in vitro models. This multifunctional liposome exhibits superior hypoxic anti-tumour effects and holds the potential to reduce side effects associated with untargeted therapy. Fluorescence of the constituent ZnPc and folate-receptor targeting could enable tracking and permit spatiotemporal regulation for improved cancer treatment.

A novel Sn(IV) meso-tetraacenaphthylporphyrin (SnTAcP) has been synthesized and characterized. SnTAcP was complexed with methyl-[Formula: see text]-cyclodextrin (m[Formula: see text]-CD), a nanocarrier that enhances water solubility, and the complexes were evaluated as PDT agents using MCF-7 breast cancer cells. A relatively low singlet oxygen quantum yield value of 0.36 was obtained in DMF, and the lowest energy Q band lies at 608 nm on the edge of the therapeutic window. SnTAcP was found to be non-toxic in the dark and phototoxic towards MCF-7 breast cancer cells with a half-maximal inhibitory concentration (IC[Formula: see text] value of 11 ± 1.1 [Formula: see text]g · mL[Formula: see text] after 30 min of irradiation with a 625 nm Thorlabs LED that provides a dose of 432 J · cm[Formula: see text]. A higher IC[Formula: see text] value of 21 ± 1.1 [Formula: see text]g · mL-1 was obtained for the m[Formula: see text]-CD inclusion complex of SnTAcP.

Novel chloroindium(III) complexes of tetra(4-methylthiophenyl)porphyrin (2a) and tetra-2-thienylporphyrin (2b) dyes have been synthesized and characterized. The main goal of the project was to identify fully symmetric porphyrin dyes with Q-band regions that lie partially in the therapeutic window that are suitable for use in photodynamic therapy (PDT). 2a and 2b were found to have fluorescence quantum yield values [Formula: see text] 0.01 and moderately high singlet oxygen quantum yields (0.54−0.73) due to heavy atom effects associated with the sulfur and indium atoms. The dark toxicity and PDT activity against epithelial breast cancer cells (MCF-7) were investigated over a dose range of 3.0−40 [Formula: see text]g [Formula: see text] mL[Formula: see text]. The in vitro dark cytotoxicity of 2a is significantly lower than that of 2b at [Formula: see text] 40 [Formula: see text]g [Formula: see text] mL[Formula: see text]. 2a was conjugated with gold nanoparticles (AuNPs) to form a nanoconjugate (2a-AuNPs), which exhibited a higher singlet oxygen quantum yield ([Formula: see text] value and PDT activity than was observed for 2a alone. The results suggest that the AuNPs nanoconjugates of readily synthesized fully symmetric porphyrin dyes are potentially suitable for PDT applications, if meso-aryl substituents that provide scope for nanoparticle conjugation can be introduced that shift the Q bands into the therapeutic window.

The synthesis, photophysicochemical and photodynamic therapy (PDT) activity of quaternary benzothiazole substituted zinc phthalocyanine (2, containing two charges, and 3, containing four charges) are reported in this work. Furthermore, the activity of the synthesized complex was compared to non-quaternary derivative (1). Higher triplet and singlet oxygen quantum yields of 0.92 and 0.85, respectively, for quaternized complexes 2 and 3 compared to complex 1 alone. Complexes 1, 2 and 3 showed relatively no dark toxicity against the epithelial breast cancer cells with cell survival of above 90 ± 3%. The quaternary derivatives (2 and 3) showed superior PDT activity with 30% or less of viable cells at concentration of 50.0 μg/mL in comparison to complex 1 alone which further lay credence to the importance of quaternization in the enhancement of PDT activity.

A 2,6-dibrominated 3,5-distyryl boron-dipyrromethene (BODIPY) dye with a moderately high singlet oxygen quantum yield was encapsulated in Pluronic® F-127 micelles, and its dark cytotoxicity and pho...

Metal free, Zn and ClGa containing carboxyphenoxy and phenoxy groups (complexes 1) and pyrene groups (complexes 2) were synthesized and embedded into Pluronic F127 micelles (represented as F127). Dark toxicity and photodynamic therapy activities of the embedded porphyrins were successfully studied on MCF-7 breast cancer cells. Dark toxicity showed more than 80% cell viability for all complexes. It was found that 1-Zn + F127 showed better photodynamic therapy activity compared to 1-H2 + F127, and 1-ClGa + F127, corresponding to the high partition coefficient for the Zn porphyrin derivatives. The same applies to 2-Zn + F127 compared to 2-H2 + F127, 2-ClGa + F127. 1-ClGa and 1-Zn were also linked to Pluronic F127 silica nanoparticles. PDT activities for embedded 1-ClGa + F127 and 1-Zn + F127 were much higher than when linked to Pluronic silica nanoparticles (PluS NPs), showing the importance of loading of porphyrins into Pluronic F127 as a drug delivering agent rather than linking. PDT studies at the highest concentration of 60 µg/ml showed decrease in cell viability down to 15.9% for 2-Zn + F127. The Kp was determined in biphasic octanol and water system.

Novel zinc(II) 3-(4-((3,17,23-tris(4-(benzo[d]thiazol-2-yl)phenoxy)phthalocyanine-9-yl)oxy)phenyl)propanoic acid (complex 3) was synthesised. Complex 3 was subsequently reacted with gold nanoparticles (AuNPs), chitosan (CT) and a gold–chitosan (AuCT) hybrid to form 3-AuNPs, 3-CT and 3-AuCT, respectively. The conjugates afforded a decrease in fluorescence quantum yield with a corresponding increase in the triplet and singlet quantum yields compared to complex 3. The in vitro dark cytotoxicity and photodynamic therapy activity (PDT) of complex 3 and 3-AuCT composites were investigated against epithelial breast cancer cells (MCF-7) with both the samples showing minimum dark cytotoxicity. They both accounted for a cell viability of ≥90% at a concentration of ≤59.2 μg mL−1. 3-AuCT showed better PDT activity (compared to 3 alone) with less than 50% viable cells at a concentration of ≥29.6 μg mL−1 making it potentially applicable for PDT. On the other hand, AuCT displayed some activity against cancer cells, probably due to photothermal activity since gold is a light absorber, however it had more than 50% viable cells at a concentration of ≤59.2 μg mL−1.

The efficacy of photodynamic therapy (PDT) is often limited by the poor bio-distributive properties of conventional photosensitizers and the local hypoxic microenvironment that characterises most solid tumours. Herein, a novel in situ oxygenic lipid formulation for photodynamic therapy (PDT) is reported. Such a hybrid was synthesized by adsorbing bimetallic nanozyme, MnO2@PtNPs (NPs = nanoparticles) onto graphene quantum dots (GQDs) – zinc (II) phthalocyanine conjugates, followed by liposomal encapsulation, affording it enhanced water solubility. The MnO2@PtNPs, which are is shown to possess excellent catalase-like properties surpassing that of MnO2 or PtNPs alone, serves to catalyze H2O2 to O2, while the zinc (II) phthalocyanine (1) serves to transform the formed oxygen to generate cytotoxic singlet oxygen immediately. We show that by combining each function of the respective building blocks, the as-synthesized 1-GQDs-MnO2@PtNPs-liposomes not only maintains the properties of oxygen supplementation through H2O2 catalysis but also displays cooperative properties for enhanced singlet oxygen production. Consequently, a remarkably improved PDT efficacy was observed for 1-GQDs-MnO2@PtNPs-liposomes in both normoxia and hypoxia. These results demonstrate the potential applicability of such nanozyme constituted 1-GQDs-MnO2@PtNPs-liposomes for achieving tumour treatment in hypoxic conditions by PDT.

Zinc monocarboxyphenoxy phthalocyanine (ZnMCPPc) was linked to human serum albumin (HSA) and chitosan via amide bond formation. The photophysical behavior and photodynamic therapy (PDT) activity (against human breast adenocarcinoma cell line (MCF-7 cells) of ZnMCPPc alone and its conjugates were investigated. The conjugates showed improved fluorescence, triplet and singlet oxygen quantum yields when compared to ZnMCPPc alone. The in vitro dark cytotoxicity and PDT studies were carried out at a dose of 3.6μg/mL to 57.1μg/mL. The in vitro dark cytotoxicity studies of ZnMCPPc showed cell viability50% at 28.6μg/mL and 57.1μg/mL, while the conjugates showed50% in all their tested concentrations (3.6 to 57.1) μg/mL. Thus, conjugation of ZnMCPPc to HSA and chitosan improves its dark cytotoxicity, an important criteria for molecules meant for photodynamic therapy. Complex 1 showed the most efficacious PDT activity with cell viability50% at concentration range of (14.3 to 57.1) μg/mL in comparison to the conjugates which only showed50% cell viability at 28.6μg/mL and 57.1μg/mL for 1-HSA and 57.1μg/mL for 1-Chitosan.

Bioprinting advances have revolutionised drug discovery and are set to disrupt biomedical research and medical application through the development of reproducible, fine-tuned functional 3D tissues and, eventually, whole organs. This intersectional bottom-up approach of additive manufacturing requires collaboration between tissue engineers, materials chemists, software and electrical engineers and medical practitioners for the software, hardware and wetware required by this disruptive technology. This review provides a current overview of the state of the art of bioprinting and the biomaterials/bioinks required, as well as the challenges and prospects for medical application in South Africa.

Aminopropyl triethoxysilane functionalized core SiO2 and core/shell ZnO/SiO2 nanoparticles (NP) were covalently linked to zinc monocarboxyphenoxy phthalocyanine (ZnMCPPc, complex 1) via amide bond formation. The photophysicochemical behavior, in vitro dark viability and photodynamic therapy (PDT) activity against human breast adenocarcinoma cell line (MCF-7 cells) of the conjugates were studied. The nanoconjugates showed enhanced photophysicochemical behavior as compared to complex 1 alone. Complex 1 showed higher dark toxicity against MCF-7 cells when compared to the conjugates. In the dark, complex 1 accounted for less than 50% viable cells at 28.6 μg/mL and 57.1 μg/mL compared to the conjugates which accounted for more than 50% cell viability at these concentrations. The in vitro dark viability and PDT activity of complex 1 was reduced in the presence of these nanoparticles.

5, 10, 15, Triphenyl-20-p-phenoxy benzoic acid porphyrins (P) containing Zn (ZnP), Ga (GaP), and Si (SiP) were synthesized and conjugated to pluronic-silica (PluS) nanoparticles (NPs) where the fluorescence and singlet oxygen generating behavior of the porphyrins were investigated. The highest singlet oxygen quantum yield (ΦΔ) was obtained for ZnP. When the porphyrins were conjugated to the PluS NPs, the ΦΔ was quenched and fluorescence was enhanced. The pore size of the NPs upon conjugation decreased from 18.9 nm for PluS NPs to 2.4 nm (for ZnP as an example) as determined by applying the Brunauer–Emmett–Teller method. The porphyrin complexes and their conjugates were tested for their photodynamic therapy (PDT) activity on MCF-7 breast cancer cells. It was found that ZnP and its conjugate showed the highest PDT activity. The p > 0.05 indicated that ZnP is significantly different than GaP and SiP.

The synthesis, photophysicochemical properties and photodynamic activity (PDT) of tetra-pyridyloxy (1,2) and benzothiazole (3, 4) substituted indium (III) (1,3) and zinc (2, 4) phthalocyanines (Pcs) and their incorporation into Pluronic® F127 and Pluronic L121/F127 mixed micelles (the latter for 3 and 4 only) are presented in this study. The InPcs exhibited higher singlet oxygen (ΦΔ) at 0.76 and 0.68 compared to the ZnPc’s at 0.47 and 0.44 in dimethyl sulfoxide. The ΦΔ values in the presence of Pluronic® F127 and in water, were 0.39 and 0.42 for InPcs and 0.23 and 0.37 for ZnPc. The ΦΔ values in the presence of Pluronic F127/L121 mixed micelles for complex 3 and 4 were 0.51 and 0.29 in water. The Kp was determined using the water and octanol system. InPcs had larger Kp values suggesting that they are more likely to be taken up by the cancer cells hence they showed better PDT activity.

Metabolic remodelling of the tumour microenvironment is a major mechanism by which cancer cells survive and resist treatment. The pro-oncogenic inflammatory cascade released by adipose tissue promotes oncogenic transformation, proliferation, angiogenesis, metastasis and evasion of apoptosis. STAT3 has emerged as an important mediator of metabolic remodelling. As a downstream effector of adipocytokines and cytokines, its canonical and non-canonical activities affect mitochondrial functioning and cancer metabolism. In this review, we examine the central role played by the crosstalk between the transcriptional and mitochondrial roles of STAT3 to promote survival and further oncogenesis within the tumour microenvironment with a particular focus on adipose-breast cancer interactions.

β-Cells contain a prominent endoplasmic reticulum (ER), disrupting ER homeostasis and function, activating the unfolded protein response (UPR). Currently, no direct protocols measure the UPR initiation. Current methods to measure ER stress include the quantification of nitric oxide (NO) (indirect method), Western blotting, and qRT-PCR of downstream components. However, these methods do not account for the overlap with mitochondrial dysfunction. In this study, INS-1E cells were exposed to proinflammatory cytokines to induce ER stress, as determined using NO, thioflavin T (ThT) binding, and β-cell functionality (insulin production). ER stress was confirmed through the upregulation of CHOP. Cell viability was monitored using MTT, sulforhodamine B, and the xCELLigence system. Morphological changes were monitored using electron microscopy. IL-1β exposure-induced β-cell stress after 4 H, decreased insulin levels, and increased thioflavin binding were noted. Increased NO production was only detected after 10 H, highlighting its lack of sensitivity, and the need for a continuous, selective, rapid, convenient, and economical detection method for early onset of ER stress. Standard methods (MTT and NO) failed to detect early ER stress. The xCELLigence coupled with a functional assay such as the detection of insulin levels or ThT are better predictors of ER stress in INS-1E cells.

Sn(iv) porphyrins ([Sn(iv)TTP(3PyO)(2)] (5) and [Sn(iv)TPP(3PyO)(2)] (6) [tetrathienylporphyrin (TTP), tetraphenylporphyrin (TPP), and pyridyloxy (PyO)]) were prepared and characterized and their photocytotoxicity upon irradiation with 625 nm light has been studied. The presence of the 3PyO axial ligands was found to limit the aggregation and enhance the solubility of 5 and 6 in DMF/H(2)O (1 : 1). The photophysical properties and photodynamic therapy (PDT) activity of the meso-2-thienyl and meso-phenyl-substituted Sn(iv) porphyrins are compared. 5 and 6 were found to be photocytotoxic in MCF-7 cancer cells when irradiated with a Thorlabs M625L3 LED at 625 nm but remained nontoxic in the dark. The PDT activity of Sn(iv) meso-tetra-2-thienylporphyrin 5 was found to be significantly enhanced relative to its analogous tetraphenylporphyrin 6. There is a marked red-shift of the Q(00) band of 5 into the therapeutic window due to the meso-2-thienyl rings, and 5 has an unusually high singlet oxygen quantum yield value of 0.83 in DMF. The results demonstrate that readily synthesized axially ligated Sn(iv) meso-arylporphyrins are potentially suitable for use as singlet oxygen photosensitizers in biomedical applications and merit further in depth investigation in this context.

In this work, the photodynamic therapy (PDT) activities (using human carcinoma adherent MCF-7 cells) of zinc phthalocyanine derivatives: complexes 1 (Zn mono cinnamic acid phthalocyanine) and 2 (zinc mono carboxyphenoxy phthalocyanine) when covalently linked to folic acid (FA) and amine functionalized magnetic nanoparticles (AMNPs) are reported. The covalent linkage of asymmetric zinc cinnamic acid Pc (1) to FA (1-FA) through an amide bond is reported for the first time. Complex 1 is insoluble in water, but upon linkage to FA, (to form 1-FA) the molecule become water soluble, hence the UV–Vis spectrum and singlet oxygen quantum yield for 1-FA were also done in water since water solubility is essential for biological applications. The reported 2-FA is also water soluble. Linking complexes 1 and 2 to FA and AMNPs decreased the dark toxicity of 1 and 2 on MCF-7 cells. Pc-FA (1-FA and 2-FA) conjugates had better singlet oxygen quantum yields (Φ∆) in DMSO as compared to Pc-AMNPs (1-AMNPs and 2-AMNPs). The water- soluble 1-FA and 2-FA also achieved a better photodynamic therapy (PDT) activity as compared to 1-AMNPs and 2-AMNPs. Folic acid targeting on the tumor cells may have also facilitated better bioavailability of 1-FA and 2-FA and improved PDT activity on MCF-7 cells over AMNPs carriers.

The syntheses of two zinc(II) phthalocyanines (ZnPcs) having either imidazole (ZnPc 1) or pyridiloxy (ZnPc 2) moieties as their macrocycle substituents are reported. Quaternization of the ZnPcs with methyl iodide afforded water soluble cationic phthalocyanines. The photophysical, photochemical properties and photodynamic therapy (PDT) activity of the ZnPcs were studied in solution. The fluorescence quantum yield and lifetime of ZnPc 1 were higher as compared to ZnPc 2. ZnPc 2 afforded higher triplet state (ΦT) and singlet oxygen quantum yields (ΦΔ) in comparison to ZnPc 1. The PDT activity of ZnPcs was investigated against human breast adenocarcinoma cells (MCF–7). The two compounds afforded a very minimal in vitro dark cytotoxicity with 85% viable cells at concentration ≤80 μM. On irradiation of the cells having the ZnPcs, ≥50% cell death was recorded for ZnPc 1 which was also evidenced by the cells photo–micrograph.

The present study shows improved photodynamic therapy (PDT) effect of zinc mono carboxy phenoxy phthalocyanine (ZnMCPPc ( 1 )) upon conjugation to spermine (via amide bond) as a targeting molecule on MCF-7 breast cancer cells. The ZnMCPPc-spermine ( 2 ) conjugate was adsorbed onto single walled carbon nanotubes (represented as ZnMCPPc-spermine-SWCNT ( 3 )). There was no change in the fluorescence quantum yield of complex 1 following formation of 2 or 3 . Complexes 2 and 3 showed improved photophysical properties; with over 50% increases in triplet and singlet oxygen quantum yields compared to 1 . Complexes 1 , 2 and 3 were relatively not toxic to MCF-7 cancer cells when incubated with 5–40 μM of each complex for 24 h in the dark. The PDT results showed that at 40 μM complex 1 resulted in only 64% decrease in cell viability, while 2 and 3 improved the PDT effect of 1 to 97% and 95% decrease in cell viability at 40 μM respectively.

Heat shock protein 70 (Hsp70) is a molecular chaperone that plays an important role in cellular proteostasis. Hsp70s are also implicated in the survival and pathogenicity of malaria parasites. The main agent of malaria, Plasmodium falciparum, expresses six Hsp70s. Of these, two (PfHsp70-1 and PfHsp70-z) localize to the parasite cytosol. Previously conducted gene knockout studies suggested that PfHsp70-z is essential, and it has been demonstrated that small-molecule inhibitors targeting PfHsp70-1 cause parasite death. For this reason, both PfHsp70-1 and PfHsp70-z are potential antimalarial targets. Two cyclic lipopeptides, colistin and polymyxin B (PMB), have been shown to bind another heat shock protein, Hsp90, inhibiting its chaperone function. In the current study, we investigated the effect of PMB on the structure-function features of PfHsp70-1 and PfHsp70-z. Using surface plasmon resonance analysis, we observed that PMB directly interacts with both PfHsp70-1 and PfHsp70-z. In addition, using circular dichroism spectrometric analysis combined with tryptophan fluorescence measurements, we observed that PMB modulated the secondary and tertiary structures of Hsp70. Furthermore, PMB inhibited the basal ATPase activity and chaperone function of the two Hsp70s. Our findings suggest that PMB associates with Hsp70 to inhibit its function. In light of the central role of Hsp70 in cellular proteostasis and its essential role in the development of malaria parasites in particular, our findings expand the library of small-molecule inhibitors that target this medically important class of molecular chaperones.

The synthesis, photophysicochemical properties and photodynamic activity (PDT) of 4(4-phenyldiazenyl) phenoxy substituted indium (III) (InPc) and zinc (ZnPc) phthalocyanines when alone or incorporated into Pluronic® F127 micelles are presented in this study. The InPc exhibited higher singlet oxygen (ΦΔ) at 0.47 compared to the ZnPc at 0.20 in dimethylsulfoxide. The ΦΔ values in the presence of Pluronic® F127 and in water, were 0.32 for InPc and 0.09 for ZnPc. The triplet quantum yields (ΦT) were 0.92 for InPc and 0.32 for ZnPc in DMSO. The PDT activity followed the same trend as the singlet oxygen quantum yields. At the highest concentration, InPc in Pluronic® F127 gave 22% cell viability compared to 34% for complex ZnPc. The partition coefficient Kp values were determined using the water and octanol system. InPc had a larger Kp suggesting that it is more likely to be taken up by the cancer cells, hence it exhibited better PDT activity.

Heat shock proteins (Hsps) play an important role in the development and pathogenicity of malaria parasites. One of the most prominent functions of Hsps is to facilitate the folding of other proteins. Hsps are thought to play a crucial role when malaria parasites invade their host cells and during their subsequent development in hepatocytes and red blood cells. It is thought that Hsps maintain proteostasis under the unfavourable conditions that malaria parasites encounter in the host environment. Although heat shock protein 70 (Hsp70) is capable of independent folding of some proteins, its functional cooperation with heat shock protein 90 (Hsp90) facilitates folding of some proteins such as kinases and steroid hormone receptors into their fully functional forms. The cooperation of Hsp70 and Hsp90 occurs through an adaptor protein called Hsp70-Hsp90 organising protein (Hop). We previously characterised the Hop protein from Plasmodium falciparum (PfHop). We observed that the protein co-localised with the cytosol-localised chaperones, PfHsp70-1 and PfHsp90 at the blood stages of the malaria parasite. In the current study, we demonstrated that PfHop is a stress-inducible protein. We further explored the direct interaction between PfHop and PfHsp70-1 using far Western and surface plasmon resonance (SPR) analyses. The interaction of the two proteins was further validated by co-immunoprecipitation studies. We observed that PfHop and PfHsp70-1 associate in the absence and presence of either ATP or ADP. However, ADP appears to promote the association of the two proteins better than ATP. In addition, we investigated the specific interaction between PfHop TPR subdomains and PfHsp70-1/ PfHsp90, using a split-GFP approach. This method allowed us to observe that TPR1 and TPR2B subdomains of PfHop bind preferentially to the C-terminus of PfHsp70-1 compared to PfHsp90. Conversely, the TPR2A motif preferentially interacted with the C-terminus of PfHsp90. Finally, we observed that recombinant PfHop occasionally eluted as a protein species of twice its predicted size, suggesting that it may occur as a dimer. We conducted SPR analysis which suggested that PfHop is capable of self-association in presence or absence of ATP/ADP. Overall, our findings suggest that PfHop is a stress-inducible protein that directly associates with PfHsp70-1 and PfHsp90. In addition, the protein is capable of self-association. The findings suggest that PfHop serves as a module that brings these two prominent chaperones (PfHsp70-1 and PfHsp90) into a functional complex. Since PfHsp70-1 and PfHsp90 are essential for parasite growth, findings from this study are important towards the development of possible antimalarial inhibitors targeting the cooperation of these two chaperones.

Heat shock protein 90 (Hsp90) has been identified in the extracellular space and has been shown to chaperone a finite number of extracellular proteins involved in cell migration and invasion. We used chemical cross-linking and immunoprecipitation followed by tandem mass spectrometry (MS/MS) to isolate a complex containing Hsp90 and the matrix protein fibronectin (FN) from breast cancer cells. Further analysis showed direct binding of Hsp90 to FN using an in vitro co-immunoprecipitation assay, a solid phase binding assay and surface plasmon resonance (SPR) spectroscopy. Confocal microscopy showed regions of co-localisation of Hsp90 and FN in breast cancer cell lines. Exogenous Hsp90β was shown to increase the formation of extracellular FN matrix in the Hs578T cell line, whilst knockdown or inhibition of Hsp90 led to a reduction in the levels of both soluble and insoluble FN and could be partially rescued by addition of exogenous Hsp90β. Treatment of cells with novobiocin led to internalization of FN into vesicles that were positive for the presence of the lysosomal marker, LAMP-1. Taken together, the direct interaction between FN and Hsp90, as well as the decreased levels of both soluble and insoluble FN upon Hsp90 inhibition or knockdown, suggested that FN may be a new client protein for Hsp90 and that Hsp90 was involved in FN matrix assembly and/or stability. The identification of FN as a putative client protein of Hsp90 suggests a role for Hsp90 in FN matrix stability, which is important for a number of fundamental cellular processes including embryogenesis, wound healing, cell migration and metastasis.

Real-time analysis offers multiple benefits over traditional end point assays. Here, we present a method of monitoring the optimisation of the growth and differentiation of murine 3T3-L1 preadipocytes to adipocytes using the commercially available ACEA xCELLigence Real-Time Cell Analyser Single Plate (RTCA SP) system. Our findings indicate that the ACEA xCELLigence RTCA SP can reproducibly monitor the primary morphological changes in pre- and post-confluent 3T3-L1 fibroblasts induced to differentiate using insulin, dexamethasone, 3-isobutyl-1-methylxanthine and rosiglitazone; and may be a viable primary method of screening compounds for adipogenic factors.