Identification and quantification of 19 pharmaceutical active compounds and metabolites in hospital wastewater in Cameroon using LC/QQQ and LC/Q-TOF.

Human pharmaceutical residues are a serious environmental concern. They have been reported to have eco, geno, and human toxic effects, and thus their importance as micropollutants cannot be ignored. These have been studied extensively in Europe and North America. However, African countries are still lagging behind in research on these micropollutants. In this study, the wastewaters of the University Teaching Hospital of Yaoundé (UTHY) were screened for the presence of active pharmaceutical ingredients and their metabolites. The screening was carried out using two methods: high-performance liquid chromatography coupled to a triple quadrupole analyzer (LC/QQQ) and high-performance coupled to a mass spectrometer with a time of flight analyzer (LC/Q-TOF). A total of 19 active pharmaceutical ingredients and metabolites were identified and quantified. The compounds identified include paracetamol (211.93 µg/L), ibuprofen (141 µg/L), tramadol (76 µg/L), O-demethyltramadol (141 µg/L), erythromycinanhydrate (7 µg/L), ciprofloxacin (24 µg/L), clarinthromycine (0.088 µg/L), azitromycine (0.39 µg/L), sulfamethoxazole 0.16 µg/L), trimetoprime (0.27 µg/L), caffeine (5.8 µg/L), carnamaeepine (0.94 µg/L), atenolol (0.43 µg/L), propranolol (0.3 µg/L), cimetidine (34 µg/L), hydroxy omeprazole (5 µg/L), diphenhydramine (0.38 µg/L), metformine (154 µg/L), and sucralose (13.07 µg/L).

Na(+)/Ca (2+) exchange and the plasma membrane Ca(2+)-ATPase in ß-cell function and diabetes.

The rat pancreatic ß-cell expresses two splice variants of the Na+/Ca(2+) exchanger 1 (NCX1) and six splice variants of the plasma membrane Ca(2+)-ATPase (PMCA). In the ß-cell, Na(+)/Ca(2+) exchange displays a high capacity, contributes to both Ca(2+) outflow and influx and participates to the control of insulin release. Gain of function studies show that overexpression of NCX1 or PMCA2 leads to endoplasmic reticulum (ER) Ca(2+) depletion with subsequent ER stress, decrease in ß-cell proliferation and ß-cell death by apoptosis. Interestingly, chronic exposure to cytokines or high free fatty acids concentration also induces ER Ca(2+) depletion and ß-cell death in diabetes. Loss of function studies shows, on the contrary, that heterozygous inactivation of NCX1 (Ncx1 ( +/- )) leads to an increase in ß-cell function (insulin production and release) and a fivefold increase in both ß-cell mass and proliferation. The mutation also increases ß-cell resistance to hypoxia, and Ncx1 ( +/- ) islets show a four to seven times higher rate of diabetes cure than Ncx1 ( +/+ ) islets when transplanted in diabetic animals. Thus, downregulation of the Na(+)/Ca(2+) exchanger leads to various changes in ß-cell function that are opposite to the major abnormalities seen in diabetes. In addition, the ß-cell, which is an excitable cell, includes the mutually exclusive exon B in the alternative splicing region of NCX1, which confers a high sensitivity of its NCX splice variants (NCX1.3 & 1.7) to the inhibitory action of compounds like KB-R7943. This provides a unique model for the prevention and treatment of ß-cell dysfunction in diabetes and following islet transplantation.

Plasma membrane Ca2+-ATPase overexpression depletes both mitochondrial and endoplasmic reticulum Ca2+ stores and triggers apoptosis in insulin-secreting BRIN-BD11 cells.

Ca(2+) may trigger apoptosis in ß-cells. Hence, the control of intracellular Ca(2+) may represent a potential approach to prevent ß-cell apoptosis in diabetes. Our objective was to investigate the effect and mechanism of action of plasma membrane Ca(2+)-ATPase (PMCA) overexpression on Ca(2+)-regulated apoptosis in clonal ß-cells. Clonal ß-cells (BRIN-BD11) were examined for the effect of PMCA overexpression on cytosolic and mitochondrial [Ca(2+)] using a combination of aequorins with different Ca(2+) affinities and on the ER and mitochondrial pathways of apoptosis. ß-cell stimulation generated microdomains of high [Ca(2+)] in the cytosol and subcellular heterogeneities in [Ca(2+)] among mitochondria. Overexpression of PMCA decreased [Ca(2+)] in the cytosol, the ER, and the mitochondria and activated the IRE1α-XBP1s but inhibited the PRKR-like ER kinase-eIF2α and the ATF6-BiP pathways of the ER-unfolded protein response. Increased Bax/Bcl-2 expression ratio was observed in PMCA overexpressing ß-cells. This was followed by Bax translocation to the mitochondria with subsequent cytochrome c release, opening of the permeability transition pore, and apoptosis. In conclusion, clonal ß-cell stimulation generates microdomains of high [Ca(2+)] in the cytosol and subcellular heterogeneities in [Ca(2+)] among mitochondria. PMCA overexpression depletes intracellular [Ca(2+)] stores and, despite a decrease in mitochondrial [Ca(2+)], induces apoptosis through the mitochondrial pathway. These data open the way to new strategies to control cellular Ca(2+) homeostasis that could decrease ß-cell apoptosis in diabetes.

Intermittent fasting modulation of the diabetic syndrome in streptozotocin-injected rats.

This study investigates the effects of intermittent overnight fasting in streptozotocin-induced diabetic rats (STZ rats). Over 30 days, groups of 5-6 control or STZ rats were allowed free food access, starved overnight, or exposed to a restricted food supply comparable to that ingested by the intermittently fasting animals. Intermittent fasting improved glucose tolerance, increased plasma insulin, and lowered Homeostatis Model Assessment index. Caloric restriction failed to cause such beneficial effects. The ß-cell mass, as well as individual ß-cell and islet area, was higher in intermittently fasting than in nonfasting STZ rats, whilst the percentage of apoptotic ß-cells appeared lower in the former than latter STZ rats. In the calorie-restricted STZ rats, comparable findings were restricted to individual islet area and percentage of apoptotic cells. Hence, it is proposed that intermittent fasting could represent a possible approach to prevent or minimize disturbances of glucose homeostasis in human subjects.

Heterozygous inactivation of the Na/Ca exchanger increases glucose-induced insulin release, ß-cell proliferation, and mass.

OBJECTIVE: We have previously shown that overexpression of the Na-Ca exchanger (NCX1), a protein responsible for Ca(2+) extrusion from cells, increases ß-cell programmed cell death (apoptosis) and reduces ß-cell proliferation. To further characterize the role of NCX1 in ß-cells under in vivo conditions, we developed and characterized mice deficient for NCX1. RESEARCH DESIGN AND METHODS: Biologic and morphologic methods (Ca(2+) imaging, Ca(2+) uptake, glucose metabolism, insulin release, and point counting morphometry) were used to assess ß-cell function in vitro. Blood glucose and insulin levels were measured to assess glucose metabolism and insulin sensitivity in vivo. Islets were transplanted under the kidney capsule to assess their performance to revert diabetes in alloxan-diabetic mice. RESULTS: Heterozygous inactivation of Ncx1 in mice induced an increase in glucose-induced insulin release, with a major enhancement of its first and second phase. This was paralleled by an increase in ß-cell proliferation and mass. The mutation also increased ß-cell insulin content, proinsulin immunostaining, glucose-induced Ca(2+) uptake, and ß-cell resistance to hypoxia. In addition, Ncx1(+/-) islets showed a two- to four-times higher rate of diabetes cure than Ncx1(+/+) islets when transplanted into diabetic animals. CONCLUSIONS: Downregulation of the Na/Ca exchanger leads to an increase in ß-cell function, proliferation, mass, and resistance to physiologic stress, namely to various changes in ß-cell function that are opposite to the major abnormalities seen in type 2 diabetes. This provides a unique model for the prevention and treatment of ß-cell dysfunction in type 2 diabetes and after islet transplantation.