Inhibitory Effect Of Chloroquine Research Articles

Potential i-Nos/Arg-1 Switch with NLRP3 and Parasitic Load Down Regulation in Experimental Schistosoma mansoni Infection via Chloroquine Repurposing.

In previous studies, the inhibitory effect of chloroquine on NLRP3 inflammasome and heme production was documented. This may be employed as a double-bladed sword in schistosomiasis (anti-inflammatory and parasiticidal). In this study, chloroquine's impact on schistosomiasis mansoni was investigated. The parasitic load (worm/egg counts and reproductive capacity index [RCI]), i-Nos/Arg-1 expression, splenomegaly, hepatic insult and NLRP3-immunohistochemical expression were assessed in infected mice after receiving early and late repeated doses of chloroquine alone or dually with praziquantel. By early treatment, the least RCI was reported in dually treated mice (41.48 ± 28.58) with a significant reduction in worm/egg counts (3.50 ± 1.29/2550 ± 479.58), compared with either drug alone. A marked reduction in the splenic index was achieved by prolonged chloroquine administration (alone: 43.15 ± 5.67, dually: 36.03 ± 5.27), with significantly less fibrosis (15 ± 3.37, 14.25 ± 2.22) than after praziquantel alone (20.5 ± 2.65). Regarding inflammation, despite the praziquantel-induced significant decrease in NLRP3 expression, the inhibitory effect was marked after dual and chloroquine administration (liver: 3.13 ± 1.21/3.45 ± 1.23, spleen: 5.7 ± 1.6/4.63 ± 2.41). i-Nos RNA peaked with early/late chloroquine administration (liver: 68.53 ± 1.8/57.78 ± 7.14, spleen: 63.22 ± 2.06/62.5 ± 3.05). High i-Nos echoed with a parasiticidal and hepatoprotective effect and may indicate macrophage-1 polarisation. On the flip side, the chloroquine-induced low Arg-1 seemed to abate immune tolerance and probably macrophage-2 polarisation. Collectively, chloroquine synergised the praziquantel-schistosomicidal effect and minimised tissue inflammation, splenomegaly and hepatic fibrosis.

Deep Transfer Learning Approach for Automatic Recognition of Drug Toxicity and Inhibition of SARS-CoV-2.

Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) causes COVID-19 and is responsible for the ongoing pandemic. Screening of potential antiviral drugs against SARS-CoV-2 depend on in vitro experiments, which are based on the quantification of the virus titer. Here, we used virus-induced cytopathic effects (CPE) in brightfield microscopy of SARS-CoV-2-infected monolayers to quantify the virus titer. Images were classified using deep transfer learning (DTL) that fine-tune the last layers of a pre-trained Resnet18 (ImageNet). To exclude toxic concentrations of potential drugs, the network was expanded to include a toxic score (TOX) that detected cell death (CPETOXnet). With this analytic tool, the inhibitory effects of chloroquine, hydroxychloroquine, remdesivir, and emetine were validated. Taken together we developed a simple method and provided open access implementation to quantify SARS-CoV-2 titers and drug toxicity in experimental settings, which may be adaptable to assays with other viruses. The quantification of virus titers from brightfield images could accelerate the experimental approach for antiviral testing.

The effect of chloroquine on the TRPC1, TRPC6, and CaSR in the pulmonary artery smooth muscle cells in hypoxia-induced experimental pulmonary artery hypertension.

Pulmonary arterial hypertension (PAH) is a life-threatening disease characterized by a constant high pulmonary artery pressure and the remodeling of the vessel. Chloroquine (CLQ) has been observed to inhibit calcium influx. The aim of this study is to investigate the effect of CLQon transient receptor cationic proteins (TRPC1 and TRPC6) and extracellular calcium-sensitive receptor (CaSR) in a hypoxic PAH model. In this study, 8- to 12-week-old 32 male Wistar albino rats, weighing 200 to 300 g, were used. The rats were studied in four groups, including normoxy control, n = 8; normoxy CLQ (50 mg/kg/28 d), n = 8; hypoxia (HX; 10% oxygen/28 d) control, n = 8; and HX (10% oxygen/28 d) + CLQ (50 mg/kg), N = 8. Pulmonary arterial medial wall thickness, pulmonary arteriole wall, TRPC1, TRPC6, and CaSR expressions were evaluated by immunohistochemistry, polymerase chain reaction, and enzyme-linked immunosorbentassay methods. At the end of the experiment, a statistically significant increase in the medial wall thickness was observed in the hypoxic group as compared with the control group. However, in the HX + CLQ group, there was a statistically significant decrease in the vessel medial wall as compared with the HX group. In the TRPC1-, TRPC6-, and CaSR-immunopositive cell numbers, messenger RNA expressions and biochemical results showed an increase in the HX group, whereas they were decreased in the HX + CLQ group. The inhibitory effect of CLQ on calcium receptors in arterioles was observed in PAH.

Potential Chronotherapeutic Optimization of Antimalarials in Systemic Lupus Erythematosus: Is Toll-Like Receptor 9 Expression Dependent on the Circadian Cycle in Humans?

Toll-like receptor 9 (TLR9) belongs to the group of endosomal receptors of the innate immune system with the ability to recognize hypomethylated CpG sequences from DNA. There is scarce information about TLR9 expression and its association with the circadian cycle (CC). Different patterns of TLR9 expression are regulated by the CC in mice, with an elevated expression at Zeitgeber time 19 (1:00 a.m.); nevertheless, we still need to corroborate this in humans. In systemic lupus erythematosus (SLE), the inhibitory effect of chloroquine (CQ) on TLR9 is limited. TLR9 activation has been associated with the presence of some autoantibodies: anti-Sm/RNP, anti-histone, anti-Ro, anti-La, and anti-double-stranded DNA. Treatment with CQ for SLE has been proven to be useful, in part by interfering with HLA-antigen coupling and with TLR9 ligand recognition. Studies have shown that TLR9 inhibitors such as antimalarial drugs are able to mask TLR9-binding sites on nucleic acids. The data presented here provide the basic information that could be useful for other clinical researchers to design studies that will have an impact in achieving a chronotherapeutic effect by defining the ideal time for CQ administration in SLE patients, consequently reducing the pathological effects that follow the activation of TLR9.

Identification of a novel autophagic inhibitor cepharanthine to enhance the anti-cancer property of dacomitinib in non-small cell lung cancer

Inhibition of autophagy is a promising strategy for non-small cell lung cancer (NSCLC) treatment, which is in the clinical trials. However, only chloroquine is used in clinic as an autophagic inhibitor and the inhibitory effect of chloroquine on autophagy is finite. Therefore, the development of an alternative autophagic inhibitor for NSCLC therapy becomes necessary. In the present study, cepharanthine (CEP), an alkaloid extracted from Stephania cepharantha Hayata, was identified as a novel autophagic inhibitor in NSCLC cells. The potential mechanism of the CEP-inhibited autophagy was by blockage of autophagosome-lysosome fusion and inhibition of lysosomal cathepsin B and cathepsin D maturation. Furthermore, we found for the first time that dacomitinib (DAC), a second-generation epidermal growth factor receptor inhibitor that in the phase III clinical trials for NSCLC treatment, induced a protective autophagy to decrease its anti-cancer effect. Combined treatment with CEP increased the anti-proliferative and apoptotic effects of DAC in vitro and enhanced the anti-cancer effect of DAC in NCI-H1975 xenograft mice. Collectively, CEP might be further developed as an autophagic inhibitor, and combined treatment of CEP and DAC could offer an effective strategy for NSCLC treatment.

Chloroquine prevents acute kidney injury induced by lipopolysaccharide in rats via inhibition of inflammatory factors

Purpose: To investigate the role of chloroquine (CQ) in lipopolysaccharide (LPS)-induced renal injury in rats. Methods: Rats were assigned to one of four groups ( n = 10). Control group was only given saline solution, whereas the model control, LPS + CQ, and LPS + yohimbine (YOH) + CQ groups were administered LPS intraperitoneally. At the end of the study, blood urea nitrogen (BUN) and creatinine (Cr) levels were determined. Results: CQ treatment significantly decreased the blood concentrations of tissue necrosis factor alpha (TNF-α), interleukin-6 (IL-6), IL-18, BUN, and Cr in the model control rats. There were also significant decreases in the levels of high mobility group protein 1 and kidney injury molecule-1 in the renal injury rats compared to the model control group. However, the inhibitory effects of CQ in the LPS-treated rats were blocked by treatment with YOH, an α-2-adrenergic receptor antagonist. Conclusions: Treatment with CQ attenuates LPS-induced renal injury by inhibiting inflammatory response. Keywords: Creatinine, Chloroquine, Inflammatory reactions, Kidney injury, Lipopolysaccharide

Chloroquine Targets Pancreatic Cancer Stem Cells via Inhibition of CXCR4 and Hedgehog Signaling

Pancreatic ductal adenocarcinoma is one of the deadliest carcinomas and is characterized by highly tumorigenic and metastatic cancer stem cells (CSC). CSCs evade available therapies, which preferentially target highly proliferative and more differentiated progenies, leaving behind CSCs as a putative source for disease relapse. Thus, to identify potentially more effective treatment regimens, we screened established and new compounds for their ability to eliminate CSCs in primary pancreatic cancer (stem) cells in vitro and corresponding patient-derived pancreatic cancer tissue xenografts in vivo. Intriguingly, we found that in vitro treatment with the antimalarial agent chloroquine significantly decreased CSCs, translating into diminished in vivo tumorigenicity and invasiveness in a large panel of pancreatic cancers. In vivo treatment in combination with gemcitabine was capable of more effectively eliminating established tumors and improved overall survival. The inhibitory effect of chloroquine was not related to inhibition of autophagy, but was due to inhibition of CXCL12/CXCR4 signaling, resulting in reduced phosphorylation of ERK and STAT3. Furthermore, chloroquine showed potent inhibition of hedgehog signaling by decreasing the production of Smoothened, translating into a significant reduction in sonic hedgehog-induced chemotaxis and downregulation of downstream targets in CSCs and the surrounding stroma. Our study demonstrates that via to date unreported effects, chloroquine is an effective adjuvant therapy to chemotherapy, offering more efficient tumor elimination and improved cure rates. Chloroquine should be further explored in the clinical setting as its success may help to more rapidly improve the poor prognosis of patients with pancreatic cancer.

Chloroquine relieves acute lung injury in rats with acute hemorrhagic necrotizing pancreatitis

SummaryThis study preliminarily investigated the mechanism by which chloroquine (CQ) relieves acute lung injury (ALI) complicated in acute hemorrhagic necrotizing pancreatitis (AHNP). Sixty male Wistar rats were randomized into sham-operated group (group A, n=10), AHNP group (group B, n=10), L-arginine-treated group (group C, n=10), L-N-nitro-L-arginine methyl ester (NAME)-treated group (group D, n=10), CQ-treated group (group E, n=10) and CQ+L-NAME-treated group (group F, n=10). TLR4 expression was measured by using real time-PCR and Western blotting respectively. The results showed that, in the group B, the expression of TLR4 and the levels of TNF-α and IL-6 in the lungs were significantly increased, and the nitric oxide (NO) concentration was reduced, as compared with those in the group A (P<0.05 or P<0.01). Lung injury was aggravated with the increased expression of TLR4. When the inhibitor and stimulator of TLR4, namely L-Arg and L-NAME, were added respectively, lung injury was correspondingly relieved or aggravated (P<0.05 or P<0.01). In the group E, TLR4 expression was substantially lower and NO concentration higher than those in the group B (P<0.05 or P<0.01). However, in the group F, NO concentration was markedly decreased, and the inhibitory effect of CQ on TLR4 expression and the relief of lung injury were weakened when compared with those in the group E (P<0.05 or P<0.01). It was concluded that TLR4 may play an important role in the pathogenesis and development of ALI complicated in AHNP. CQ could relieve ALI by decreasing the TLR4 expression and increasing the NO release.

Inhibitory effects of chloroquine on the activation of plasmacytoid dendritic cells in SIVmac239-infected Chinese rhesus macaques

It is currently widely accepted that immune activation in HIV-infected individuals leads to a severe loss of CD4⁺ T cells and the progression to AIDS. However, the underlying mechanism of this immune activation remains unclear. Experimental data suggest that the activation of plasmacytoid dendritic cells (pDCs) by plasma viremia may play a critical role in HIV-induced immune activation. In this study, we found that the level of immune activation was higher in the late phase of SIVmac239 infection compared with chronic infection, which suggests that immune activation might be related to disease progression in SIVmac239-infected non-human primate models. Our work also showed that chloroquine could effectively inhibit the activation of pDCs in vitro and in vivo. However, chloroquine treatment of SIVmac239-infected macaques had no significant influence on the Cellular composition of peripheral blood in these animals.

Inhibitory effect of chloroquine on bone resorption reveals the key role of lysosomes in osteoclast differentiation and function

The lysosome is an acidic compartment containing certain hydrolytic enzymes necessary for the intracellular digestion of macromolecules. In terms of the activity of bone-resorbing osteoclasts, the secretion of lysosomal vesicles containing protons and matrix-degrading proteinases into the resorption lacunae is essential. Chloroquine (CQ), one of the lysosomotropic agents, has an immunosuppressive effect and is used for the treatment for rheumatoid arthritis (RA) and systemic lupus erythematosus (SLE). However, the direct effect of CQ on osteoclasts has not been reported. Here, we show that CQ suppresses the bone resorbing activity of osteoclasts by inhibition of the acidification in the lysosomes, as well as osteoclast differentiation in vitro. CQ treatment ameliorates the bone loss induced by RANKL injection in mice. These results suggest that CQ has a bone-increasing effect by inhibiting osteoclast differentiation and function. In addition, a lysosomal proton pump inhibitor bafilomycin A1 also inhibits osteoclast differentiation. Thus, this study revealed the importance of the lysosomes in osteoclast differentiation and function in vivo as well as in vitro, suggesting the therapeutic efficacy of immunosuppressive CQ in osteoclast-mediated bone loss or destruction.

The Nature of the Active Ionic Species in the Inhibitory Effect of Chloroquine Against Escherichia Coli

The Nature of the Active Ionic Species in the Inhibitory Effect of Chloroquine Against <i>Escherichia Coli</i>

Synergistic inhibitory effect of chloroquine combined with topotecan on proliferation of colon cancer cells.

Objective:The lysosomotropic drug chloroquine(CQ) has been shown in clinical trials to enhance chemotherapy and radiation sensitivity.This study is designed to examine the inhibitory effects of CQ combined with topotecan(TPT)on the proliferation of colon cancer cells and elucidate the potential mechanisms.Methods:Colon cancer SW480,SW620,and LoVo cells were treated with TPT alone or TPT combined with CQ at non cytotoxic concentrations.Cell viability was examined using MTT assay.Expressions of autophagy-related proteins LC3,P62,and Beclin1 in LoVo cells were examined by Western blotting after CQ or TPT treatment,and the intracellular accumulation of YFP-LC3 dots was examined by confocal microscopy.After Beclin1 gene was silenced by small interfering RNA(siRNA) in LoVo cells,the death rate was evaluated by Trypan blue staining after TPT treatment.Results:CQ at low concentrations enhanced the inhibitory effects of TPT on the proliferation of LoVo cells.CQ blocked TPT-induced autophagy by interffering the function of autophagic lysosome.Silencing autophagy-related gene Beclin1 by siRNA also enhanced the cytotoxicity of TPT.Conclusion:CQ potentiated the cytotoxicity of TPT on colon cancer cells.The mechanism may be related with blockade of autophagy induced by TPT at non cytotoxic concentration.This implies that CQ has significant potential to be a novel chemotherapeutic enhancer for colon cancer therapy.

Inhibitory Properties of Nerve-Specific Human Glutamate Dehydrogenase Isozyme by Chloroquine

Human glutamate dehydrogenase exists in hGDH1 (housekeeping isozyme) and in hGDH2 (nerve-specific isozyme), which differ markedly in their allosteric regulation. In the nervous system, GDH is enriched in astrocytes and is important for recycling glutamate, a major excitatory neurotransmitter during neurotransmission. Chloroquine has been known to be a potent inhibitor of house-keeping GDH1 in permeabilized liver and kidney-cortex of rabbit. However, the effects of chloroquine on nerve-specific GDH2 have not been reported yet. In the present study, we have investigated the effects of chloroquine on hGDH2 at various conditions and showed that chloroquine could inhibit the activity of hGDH2 at dose-dependent manner. Studies of the chloroquine inhibition on enzyme activity revealed that hGDH2 was relatively less sensitive to chloroquine inhibition than house-keeping hGDH1. Incubation of hGDH2 was uncompetitive with respect of NADH and non-competitive with respect of 2-oxoglutarate. The inhibitory effect of chloroquine on hGDH2 was abolished, although in part, by the presence of ADP and L-leucine, whereas GTP did not change the sensitivity to chloroquine inhibition. Our results show a possibility that chloroquine may be used in regulating GDH activity and subsequently glutamate concentration in the central nervous system.

Modulation of Arginase Pathway by Chloroquine and Its Congener May Underlie the Enhanced Cell Commitment towards Erythroid Differentiation.

Arginase is a nitric oxide synthase-alternative pathway enzyme for L-arginine breakdown leading to biosynthesis of urea and L-ornithine. Evidence suggests that reduction of arginine bioavailability for NO production secondary to increased plasma arginase levels in sickle cell disease, may contribute to severe pathophysiological derangements and increase mortality. Recently, we showed that Chloroquine (CQ), an anti-malarial and anti-rheumatoid drug, displays a linear competitive mode of inhibition on sickle erythrocyte arginase (Iyamu et al., Brit J. Haematol, 2007: in press). Utilizing the K562 cell line as a model system in our present study, we obtained evidence of the anti-proliferative and differentiation effects of CQ and its analog, Hydroxychloroquine (OHCQ) at pharmacologically attainable concentrations (5–20μM). Specifically, CQ (IC50= 8.0 ±1.2 μM) and OHCQ (IC50 =17.0 ± 2.4 μM) inhibited K562 cell proliferation in a dose-dependent manner. This inhibitory effect was accompanied by enhanced commitment towards erythroid maturation as assessed by hemoglobinization (Iyamu et al., Exp Hematol, 2003); this was linked to a dose-dependent inhibition of arginase activity. To assess the role of the arginine-polyamine pathway in the induction of differentiation of K562 cells by CQ or OHCQ, cells were treated with 10 μM CQ or 20 μM OHCQ in the presence or absence of spermine (5μM) or spermidine (5 μM). Our results indicate that spermine and spermidine (intermediate products of the arginine polyamine pathway) partially reverted the effects of CQ and OHCQ on arginase activity. But this reversal did not affect the commitment of K562 cells towards erythroid maturation. We also show that CQ and OHCQ enhanced fetal hemoglobin (Hb F) synthesis by 3.4 and 3.2-fold (vs control), and maximally stimulated intracellular cGMP levels (as determined by immunosorbent assay) by 6.6- and 3.0-fold at 6hrs and 3hrs respectively. The induction of these macromolecules by CQ or OHCQ correlated with arginase inhibition. Again, the partial reversal of arginase activity by spermine or spermidine could not reduce the Hb F induction by CQ or OHCQ. This observation suggests that the commitment of cells towards erythroid maturation precedes the effects of spermine or spermidine on arginase activity. We further investigated the effects of 8-Bromo-cAMP (cAMP analog) on arginase activity in lipopolysaccharide (LPS)-stimulated K562 cells. The inhibitory effect of CQ on arginase was reversed in LPS-stimulated cells but this reversal of arginase activity was not observed in Bromo-cAMP-LPS- treated cells. Moreover, the moderate increase of arginase activity in LPS-treated cells in the presence of CQ failed to reduce the CQ-dependent enhancement of total hemoglobin and Hb F production. Indeed, the combination of 8-Bromo-cAMP with CQ in LPS-treated cells resulted in a significant inhibition (>35%) of arginase activity. This inhibitory effect was associated with a 3.9-fold increase of Hb F synthesis (vs CQ alone) as well as enhancement of total Hb production. This surprising observation demonstrates the possible involvement of the arginase pathway as well as additional, but yet unidentified mechanism(s). In conclusion, these observations that CQ and OHCQ may inhibit the initial step of the arginine-polyamine pathway, resulting in the enhancement of erythroid differentiation may provide a paradigm for targeted therapy of hemoglobinopathies.

Effect of chloroquine on strips of non-pregnant and pregnant mice uteri in-vitro

Chloroquine (CQ) remains the household drug for the treatment of malaria especially among pregnant women. However, there are reports that CQ inhibits the contractile process in non-pregnant rat's uterus. The aim of this study is to compare responses to CQ between non-pregnant and pregnant mice and identify some mechanisms involved. Experiments were carried out in non-pregnant and pregnant mice pretreated 24 hours before with 1.5 mg/kg-body weight stilboesterol given orally. Strips of uterine smooth muscle, approximately 5 mm in diameter, were mounted in a 20 ml organ bath containing De Jalon solution bubbled with a 95% O2-5% CO2 gas mixture. Responses of the strips to graded concentration of acetylcholine (ACh) (10(-9) to 10(-5) mol/L), oxytocin (OXY) (10(-5) to 10(-2) IU/ml) and CQ (10(-6) to 4 x 10(-4) mol/L) were investigated. The strips were then incubated in 4 x 10(-4) mol/L CQ for 15 mins and the cumulative dose responses for OXY were repeated. To investigate mechanism of action, the strips were incubated for 15 mins in N(w)-nitro L-arginine methyl ester (L-NAME) and the cumulative responses to CQ repeated. Each investigation was carried out in fresh tissue mounted on Grass Model FT03 force transducer coupled unto a 4-channel Grass Model 7D Polygraph. CQ (low to moderate level), ACh and OXY led to increases in contractile responses in the uteri. There were greater contractile responses in non-pregnant than pregnant mice to CQ and ACh. At high doses, CQ had an inhibitory effect on the uterine contraction. Incubating in CQ led to abolition of contractile responses to OXY and ACh. In the presence of L-NAME, inhibitory effect of CQ at high doses was attenuated in pregnant mice only. The results suggest that CQ at high doses inhibits contractile responses in non-pregnant and pregnant mice. Enhanced nitric oxide bioactivity attenuated this inhibitory effect.

Effect of chloroquine on arachidonic acid pathway in isolated mast cells

Chloroquine (CQ) interacts with the arachidonic acid (AA) pathway in many cells and tissues as a result of phospholipase A2 inhibition. Previously, we reported the inhibitory effect of CQ on stimulated secretion from isolated rat mast cells along with the inhibition of stimulated AA liberation and thromboxane B2 (TXB2) production from membrane phospholipids [1, 2]. In this study we investigated the effect of CQ on prostagiandin D2 (PGD2) generation (a major cyclooxygenase metabolite of arachidonic acid in mast cells) [3], in compound 48/80 stimulated isolated mast cells and evaluated the relationship between AA liberation, PGD2 and TXB2 generation from membrane phospholipids.

Inhibitory effect of chloroquine on the peroxidase activity of ferriprotoporphyrin IX

The effect of the antimalarial chloroquine {7-chloro-4-[4-(diethylamino)-1-methylbutylamino]quinoline} on the oxidation of 2,2′-azinobis(3-ethylbenzothiazoline-6-sulfonate)(abts) promoted by ferriprotoporphyrin IX [(3,7,12,17-tetramethyl-8,13-divinylporphyrin-2,18-dipropanoato)iron(III)] and H2O2 was studied. Under the conditions [abts][H2O2][catalyst], the kinetic results showed a first-order dependence on both the peroxide (up to 8 × 10–3 mol dm–3) and the catalyst concentration (up to 4 × 10–6 mol dm–3), although a saturation effect was observed at higher concentrations. The values of the apparent second-order rate constants in the absence and presence of chlorcquine were (1.71 ± 0.05)× 105 and (1.23 ± 0.04)× 105 dm6 mol–2 s–1 at pH 6.86, 303.1 ± 0.2 K, and I= 0.10 mol dm–3 NaCl. Therefore, the antimalarial inhibits the peroxidase activity by about 28% due the formation of a catalytically inactive complex with the haem. A first-order dependence on the concentration of abts was also verified, followed by a zero-order dependence at concentrations higher than 5 × 10–3 mol dm–3. Some EPR spin-trapping experiments, and comparative studies in the presence of bromide ions, provided evidence for the participation of analogues of the horseradish peroxidase compounds I and II rather than hydroxyl radicals, in the peroxidase activity of the haem. The results are discussed in the context of the antimalarial activity of chloroquine.

On the inhibitory effect of chloroquine on blood platelet aggregation

Chloroquine inhibited aggregation of rat blood platelets in a dose-dependent way. The inhibitory effect decreased, depending on the aggregation stimulus used, in the rank order of potency: ADP > thrombin > A23187. Effect of 1 millimolar chloroquine on A23187-stimulated aggregation was significantly decreased or completely abolished by addition of Ca 2+ ions in the concentration of 0.1 or 1 mmol/1, respectively. For thrombin-stimulated aggregation the effect of chloroquine was partially decreased by corresponding isomolar calcium; platelet aggregation induced by ADP was not changed in the presence of extracellular Ca 2+ ions. Addition of chloroquine during aggregation resulted in disaggregation of ADP-stimulated platelets and inhibition of thrombin-and A23187-induced aggregation.

Processing and characterization of the low density lipoprotein receptor in the human colonic carcinoma cell subclone HT29-18: a potential pathway for delivering therapeutic drugs and genes.

Low density lipoprotein (LDL) processing has been investigated in the subcloned human colonic carcinoma cell line HT29-18. LDL binding at 4 degrees C was a saturable process in relation to time and LDL concentration. The Kd for LDL binding was 11 micrograms/ml. ApoE-free HDL3 or acetylated LDL did not significantly compete with 125I-LDL binding, up to 500 micrograms/ml. 125I-LDL binding was decreased by 70% in HT29-18 cells preincubated for 24 hours in culture medium containing 100 micrograms/ml unlabelled LDL. Ligand blotting studies performed on HT29-18 homogenates using colloidal gold labelled LDL indicated the presence of one autoradiographic band corresponding to an apparent molecular weight of 130 kDa, which is consistent with the previously reported molecular weight of the LDL receptor in human fibroblasts. At 37 degrees C, 125I-LDL was actively internalized by HT29-18 cells and lysosomal degradation occurred as demonstrated by the inhibitory effect of chloroquine. LDL uptake and degradation by HT29-18 cells also resulted in a marked decrease in endogenous sterol synthesis. These data demonstrate that the HT29-18 human cancerous intestinal cells are able to specifically bind and internalize LDL, and that LDL processing results in down-regulation of sterol biosynthesis. Thus, intestinal epithelial cells possess specific LDL receptors that can be exploited to accomplish drug delivery and gene transfer via the receptor-mediated endocytosis pathway.

Inhibitory effects of chloroquine on rat mast cell activation

Chloroquine failed to liberate histamine in a dose-dependent manner in the absence of extracellular calcium, but it inhibited histamine liberation induced by compound 48/80.45Ca uptake and3H-arachidonic acid liberation stimulated by compound 48/80 were also significantly inhibited by chloroquine pretreatment. The inhibitory effects of chloroquine may be accounted for by its interaction with calcium movement, as well as with prostaglandin synthesis presumably by inhibiting phospholipase A2 activity in stimulated mast cells.