Mutation in Smek2 regulating hepatic glucose metabolism causes hypersarcosinemia and hyperhomocysteinemia in rats.

Suppressor of mek1 (Dictyostelium) homolog 2 (Smek2), was identified as one of the responsible genes for diet-induced hypercholesterolemia (DIHC) of exogenously hypercholesterolemic (ExHC) rats. A deletion mutation in Smek2 leads to DIHC via impaired glycolysis in the livers of ExHC rats. The intracellular role of Smek2 remains obscure. We used microarrays to investigate Smek2 functions with ExHC and ExHC.BN-Dihc2BN congenic rats that harbor a non-pathological Smek2 allele from Brown-Norway rats on an ExHC background. Microarray analysis revealed that Smek2 dysfunction leads to extremely low sarcosine dehydrogenase (Sardh) expression in the liver of ExHC rats. Sarcosine dehydrogenase demethylates sarcosine, a byproduct of homocysteine metabolism. The ExHC rats with dysfunctional Sardh developed hypersarcosinemia and homocysteinemia, a risk factor for atherosclerosis, with or without dietary cholesterol. The mRNA expression of Bhmt, a homocysteine metabolic enzyme and the hepatic content of betaine (trimethylglycine), a methyl donor for homocysteine methylation were low in ExHC rats. Results suggest that homocysteine metabolism rendered fragile by a shortage of betaine results in homocysteinemia, and that Smek2 dysfunction causes abnormalities in sarcosine and homocysteine metabolism.

Quantitative analysis of sarcosine with special emphasis on biosensors: a review.

The quantitative determination of sarcosine is of great importance in clinical chemistry, food and fermentation industries. Elevated sarcosine levels are associated with Alzheimer, dementia, prostate cancer, colorectal cancer, stomach cancer and sarcosinemia. This review summarizes the various methods for quantitative analysis of sarcosine with special emphasis on various strategies of biosensors and their analytical performance. The current bio sensing methods have overcome the drawbacks of conventional methods. Sarcosine biosensors work optimally at pH 7.0 to 8.0 in the linear range of 0.1 to 100 µM within 2 to 17 s and between 25 and 37 °C, within a limit of detection (LOD) between 0.008 and 500 mM. The formulated biosensors can be reused within a stability period of 3-180 days. Future research could be focused to modify existing sarcosine biosensors, leading to simple, reliable, and economical sensors ideally suited for point-of-care treatment. Clinical significance Elevated sarcosine levels are associated with prostate and colorectal cancer, Alzheimer, dementia, stomach cancer and sarcosinemia. Quantitative determination of sarcosine is of great importance in clinical chemistry as well as food and fermentation industries. Attempts made in development of sarcosine biosensors have been reviewed with their advantages and disadvantages, so that scientist and clinicians can improvise the methods of developing more potent sarcosine biosensor applicable in multitudinous fields. This is the first comprehensive review which compares the various immobilization methods, sensing principles, strategies used in biosensors and their analytical performance in detail.

Chemically induced acute model of sarcosinemia in wistar rats.

In the present study, we developed an acute chemically induced model of sarcosinemia in Wistar rats. Wistar rats of 7, 14 and 21 postpartum days received sarcosine intraperitoneally in doses of 0.5 mmol/Kg of body weight three time a day at intervals of 3 h. Control animals received saline solution (NaCl 0.85 g%) in the same volume (10 mL/Kg of body weight). The animals were killed after 30 min, 1, 2, 3 or 6 h after the last injection and the brain and the blood were collected for sarcosine measurement. The results showed that plasma and brain sarcosine concentrations achieved levels three to four times higher than the normal levels and decreased in a time-dependent way, achieving normal levels after 6 hours. Considering that experimental animal models are useful to investigate the pathophysiology of human disorders, our model of sarcosinemia may be useful for the research of the mechanisms of neurological dysfunction caused by high tissue sarcosine levels.

Discovery of Tricyclic Clerodane Diterpenes as Sarco/Endoplasmic Reticulum Ca(2+)-ATPase Inhibitors and Structure-Activity Relationships.

Tricyclic clerodane diterpenes (TCDs) are natural compounds that often show potent cytotoxicity for cancer cells, but their mode of action remains elusive. A computationally based similarity search (CDRUG), combined with principal component analysis (ChemGPS-NP) and docking calculations (GOLD 5.2), suggested TCDs to be inhibitors of the sarco/endoplasmic reticulum Ca(2+)-ATPase (SERCA) pump, which is also the target of the sesquiterpene lactone thapsigargin. Biochemical studies were performed with 11 TCDs on purified rabbit skeletal muscle sarcoplasmic reticulum membranes, which are highly enriched with the SERCA1a isoform. Casearborin D (2) exhibited the highest affinity, with a KD value of 2 µM and giving rise to complete inhibition of SERCA1a activity. Structure-activity relationships revealed that functionalization of two acyl side chains (R1 and R4) and the hydrophobicity imparted by the aliphatic chain at C-9, as well as a C-3,C-4 double bond, play crucial roles for inhibitory activity. Docking studies also suggested that hydrophobic interactions in the binding site, especially with Phe256 and Phe834, may be important for a strong inhibitory activity of the TCDs. In conclusion, a novel class of SERCA inhibitory compounds is presented.

SBF-1 exerts strong anticervical cancer effect through inducing endoplasmic reticulum stress-associated cell death via targeting sarco/endoplasmic reticulum Ca(2+)-ATPase 2.

Cervical cancer is one of the most common carcinomas in the genital system. In the present study, we report that SBF-1, a synthetic steroidal glycoside, has a strong antigrowth activity against human cervical cancer cells in vitro and in vivo. SBF-1 suppressed the growth, migration and colony formation of HeLa cells. In addition, severe endoplasmic reticulum (ER) stress was triggered by SBF-1, and 4-phenyl-butyric acid, a chemical chaperone, partially reversed SBF-1-induced cell death. To uncover the target protein of SBF-1, the compound was labeled with biotin. The biotin-labeled SBF-1 bound to sarco/ER Ca(2+)-ATPase 2 (SERCA2) and colocalized with SERCA2 in HeLa cells. Moreover, SBF-1 inhibited SERCA activity, depleted ER Ca2+ and increased cytosolic Ca2+ levels. 1,2-Bis(o-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid, a chelator of Ca2+, partially blocked SBF-1-induced ER stress and growth inhibition. Importantly, knockdown of SERCA2 increased the sensitivity of HeLa cells to SBF-1-induced ER stress and cell death, whereas overexpression of SERCA2 decreased this sensitivity. Furthermore, SBF-1 induced growth suppression and apoptosis in HeLa xenografts, which is closely related to the induction of ER stress and inhibition of SERCA activity. Finally, SERCA2 expression was elevated in human cervical cancer tissues (n=299) and lymph node metastasis (n=8), as compared with normal cervix tissues (n=23), with a positive correlation with clinical stages. In all, these results suggest that SBF-1 disrupts Ca2+ homeostasis and causes ER stress-associated cell death through directly binding to SERCA2 and inhibiting SERCA activity. Our findings also indicate that SERCA2 is a potential therapeutic target for human cervical cancer.

Mutations in the sarcosine dehydrogenase gene in patients with sarcosinemia.

Sarcosinemia is an autosomal recessive metabolic trait manifested by relatively high concentrations of sarcosine in blood and urine. Sarcosine is a key intermediate in 1-carbon metabolism and under normal circumstances is converted to glycine by the enzyme sarcosine dehydrogenase. We encountered six families from two different descents (French and Arab), each with at least one individual with elevated levels of sarcosine in blood and urine. Using the "candidate gene approach" we sequenced the gene encoding sarcosine dehydrogenase (SARDH), which plays an important role in the conversion of sarcosine to glycine, and found four different mutations (P287L, V71F, R723X, R514X) in three patients. In an additional patient, we found a uniparental disomy in the region of SARDH gene. In two other patients, we did not find any mutations in this gene. We have shown for the first time that mutations in the SARDH gene are associated with sarcosinemia. In addition, our results indicate that other genes are most probably involved in the pathogenesis of this condition.

A report of two families with sarcosinaemia in Hong Kong and revisiting the pathogenetic potential of hypersarcosinaemia.

INTRODUCTION: Sarcosinaemia is a rare metabolic disorder which has not been reported in Asia. CLINICAL PICTURE: The urine samples of 2 patients were screened as a routine metabolic screening offered for patients with mental retardation in our hospital. We used gas chromatography-mass spectrometry (GC-MS) which is capable of detecting abnormal pattern in amino acids and organic acids. Plasma sarcosine level was further quantified by GC-MS. The same methods were used in the investigations of asymptomatic family members. Urine examination by GC-MS revealed excessive amount of sarcosine in urine (normally undetectable) and their plasma sarcosine levels were raised. The 2 differential diagnoses of presence of sarcosine in urine--glutaric aciduria type II and folate deficiency--were ruled out by the absence of abnormal organic acids in the initial urine screen and by normal serum folate level respectively. Screening of the 2 families identified excessive sarcosine in urine in 2 siblings, one from each family. However, these 2 siblings of indexed patients thus identified have no neurological or developmental problem. CONCLUSION: Our finding was consistent with the notion that sarcosinaemia is a benign condition picked up coincidentally during screening for mental retardation.