Artemisinin resistance in P. falciparum: probing the interacting partners of Kelch13 protein in parasite.

OBJECTIVES: Artemisinin (ART) resistance in Plasmodium is threatening the artemisinin combination therapies-the first line of defence against malaria. ART resistance has been established to be mediated by the Plasmodium Kelch13 (PfK13) protein. For the crucial role of PfK13 in multiple pathways of the Plasmodium life cycle and ART resistance, it is imperative that we investigate its interacting partners. METHODS: We recombinantly expressed PfK13-p (Bric a brac/Poxvirus and zinc finger and propeller domains), generating anti-PfK13-p antibodies to perform co-immunoprecipitation assays and probed PfK13 interacting partners. Surface plasmon resonance and pull-down assays were performed to establish physical interactions of representative proteins with PfK13-p. RESULTS: The co-immunoprecipitation assays identified 17 proteins with distinct functions in the parasite life cycle- protein folding, cellular metabolism, and protein binding and invasion. In addition to the overlap with previously identified proteins, our study identified 10 unique proteins. Fructose-biphosphate aldolase and heat shock protein 70 demonstrated strong biophysical interaction with PfK13-p, with KD values of 6.6 µM and 7.6 µM, respectively. Additionally, Plasmodium merozoite surface protein 1 formed a complex with PfK13-p, which is evident from the pull-down assay. CONCLUSION: This study adds to our knowledge of the PfK13 protein in mediating ART resistance by identifying new PfK13 interacting partners. Three representative proteins-fructose-biphosphate aldolase, heat shock protein 70, and merozoite surface protein 1-demonstrated clear evidence of biophysical interactions with PfK13-p. However, elucidation of the functional relevance of these physical interactions are crucial in context of PfK13 role in ART resistance.

Ginsenoside Rg1 exerts anti­apoptotic effects on non­alcoholic fatty liver cells by downregulating the expression of SGPL1.

Non­alcoholic fatty liver disease (NAFLD) has a high incidence, and can lead to liver cirrhosis and even hepatocellular carcinoma in severe cases. To the best of our knowledge, there is currently no safe and effective treatment for the management of this disease. Ginsenoside Rg1 (Rg1) is an active monomer derived from ginseng and notoginseng. In the present study, HHL­5 hepatocytes were used to establish an in vitro cell model of NAFLD by medium­ and long­chain fat emulsion treatment, and the effects of Rg1 on adipose accumulation, apoptosis and the expression levels of apoptosis­related proteins in HHL­5 hepatocytes were examined. The results demonstrated that Rg1 inhibited the accumulation of fat in HHL­5 cells, while inhibiting apoptosis, and Rg1 downregulated the expression levels of the pro­apoptotic protein Bax and upregulated the expression levels of the anti­apoptotic protein Bcl­2, indicating that Rg1 could promote the stability or integrity of mitochondria and exert an anti­apoptotic effect by regulating Bcl­2 family proteins. In addition, Rg1 markedly downregulated the expression levels of sphingosine­1­phosphate lyase 1 (SGPL1), a key enzyme in the sphingosine signaling pathway, in HHL­5 cells with steatosis, and increased the expression levels of the downstream pro­survival signals phosphorylated (p­)Akt and p­Erk1/2. Furthermore, overexpression of SGPL1 abolished the anti­apoptotic effect of Rg1 on SGPL1­overexpressing HHL­5 cells with steatosis, and downregulated the expression levels of pro­survival proteins, such as Bcl­2, p­Akt and p­Erk1/2, whereas the expression levels of pro­apoptotic Bax were markedly increased. In conclusion, although there are some reports regarding the protective effect of Rg1 on fatty liver cells, to the best of our knowledge, the present study is the first to report that Rg1 may exert an anti­apoptotic effect on fatty liver cells by regulating SGPL1 in the sphingosine signaling pathway. Rg1 is the main component of the prescription drug Xuesaitong in China; therefore, the findings of the present study may provide a theoretical molecular basis for the use of Rg1 or Xuesaitong in the treatment of patients with NAFLD.

Amelioration of frozen gait by tandospirone, a serotonin 1A agonist, in a patient with pure akinesia developing resistance to L-threo-3,4-dihydroxyphenylserine.

A 71-year-old woman presented with severe akinesia, frozen gait, and compromised postural reflexes, without rigidity, tremor, or vertical gaze disturbance. With a working diagnosis of pure akinesia, we administered amantadine (150 mg/d) and L-threo-3,4-dihydroxyphenylserine (DOPS) (600 mg/d), which alleviated her symptoms. When frozen gait recurred 2 months later, we increased the dose of L-threo-DOPS to 900 mg/d and added levodopa (300 mg/d) combined with carbidopa, but this failed to improve the patient's symptoms. We then combined administration of tandospirone, a serotonin (5-HT) 1A agonist with L-threo-DOPS (600 mg/d), resulting in marked clinical improvement. Tandospirone is reported to activate noradrenergic neurons via the 5-HT 1A receptor, which could account for such striking improvement in a patient previously responsive to the noradrenergic precursor L-threo-DOPS given alone.

L-threo-3,4-dihydroxyphenylserine enhances the orthostatic responses of plasma renin activity and angiotensin II in multiple system atrophy.

We evaluated the effect of L-threo-3,4-dihydroxyphenylserine (L-threo-DOPS) on the response of blood pressure and vasoactive factors during head-up tilt in patients with multiple system atrophy (MSA). After the daily oral administration of 300 mg L-threo-DOPS for 2 weeks we observed a significant reduction in the decline of systolic blood pressure during 60 degrees head-up-tilt. In concordance with this result, we detected significant increases in postural changes in plasma renin activity and angiotensin II following L-threo-DOPS treatment. The enhanced responses of these vasoactive mediators by L-threo-DOPS during head-up tilt may contribute to the improvement in orthostatic hypotension in patients with MSA.

[Effects of L-threo-Dops on orthostatic hypotension in Parkinson's disease].

Effects of L-threo-Dops (Dops) administration on orthostatic hypotention were evaluated with changes in blood pressure by postural change (lying to standing position) and subjective symptoms in 15 patients of Parkinson's disease having symptoms of orthostatic hypotention. Orthostatic syncope had improved significantly (p < 0.01) after 2 and 4 weeks of administration with maintenance dose of 460 mg/day of Dops in average. In the standing-up (Schellong) test, decrease in blood pressure levels by a postural change, both with systolic and diastolic blood pressure, was significantly smaller at 3, 5 and 10 minutes after standing after 4 weeks of drug administration. Decrease in the blood pressure level immediately after standing-up improved by 10.2 +/- 4.0 for systolic and 6.5 +/- 1.8 for diastolic (mmHg, mean +/- SE) (p < 0.01). The group that showed improvement in orthostatic syncope had a significant improvement in decline in blood pressure by standing after administration of Dops, while the group without any change in severity of syncope did not show significant improvement in orthostatic hypotention.