Expanding the phenotypic and genotypic characteristics of trichohepatoenteric syndrome: a report of eight patients from five unrelated families.

BACKGROUND: Trichohepatoenteric syndrome (THES) is characterized by neonatal-onset intractable diarrhea. It often requires long-term total parenteral nutrition (TPN). In addition, other characteristic findings of the syndrome include growth retardation, facial dysmorphism, hair abnormalities, various immunological problems and other rare system findings. Two genes and their associated pathogenic variants have been associated with this syndrome: SKIC3 and SKIC2. METHODS AND RESULTS: In this case series, the clinical findings and molecular analysis results of a total of 8 patients from 5 different families who presented with persistent diarrhea and were diagnosed with THES were shared. Pathogenic variants were detected in the SKIC3 gene in 6 of our patients and in the SKIC2 gene in 2 patients. It was planned to compare the clinical findings of our patients with other patients, together with literature data, and to present yet-undefined phenotypic features that may be related to THES. In our case series, in addition to our patients with a novel variant, patient number 2 had a dual phenotype (THES and Spondyloepimetaphyseal dysplasia, sponastrime type) that has not been reported yet. Delay in gross motor skills, mild cognitive impairment, radioulnar synostosis, osteoporosis, nephropathy and cystic lesions (renal and liver) were observed as unreported phenotypic findings. CONCLUSIONS: We are expanding the clinical and molecular repertoire of the syndrome regarding patients diagnosed with THES. We recommend that the NGS (next-generation sequencing) multigene panel should be used as a diagnostic tool in cases with persistent diarrhea.

Carbamazepine-induced sperm disorders can be associated with the altered expressions of testicular KCNJ11/miR-let-7a and spermatozoal CFTR/miR-27a.

Male infertility is a global health problem, and the underlying molecular mechanisms are not clearly known. Ion channels and microRNAs (miRNAs), known to function in many vital functions in cells, have been shown to play a significant role in male infertility through changes in their expressions. The study aimed to evaluate the alterations of testicular and/or spermatozoal potassium voltage-gated channel subfamily J member 11 (KCNJ11), Cystic fibrosis transmembrane conductance regulator (CFTR), miR-let-7a and miR-27a expressions in carbamazepine-related male infertility. Here, we showed that carbamazepine reduced sperm motility, increased abnormal sperm morphology, and impaired hormonal balance as well as increased relative testis weight and decreased relative seminal vesicle weight. On the other hand, downregulated KCNJ11 and upregulated miR-let-7a expressions were determined in testis (p < .05). Also, downregulated KCNJ11 and upregulated CFTR and miR-27a expressions were found in spermatozoa (p < .05). Interestingly, altered testicular KCNJ11 and miR-let-7a expressions were correlated with decreased sperm motility and elevated sperm tail defect. Besides, spermatozoal CFTR and miR-27a expressions positively correlated with sperm tail defects. The results indicated a significant relationship between ion channel and/or miRNA expression alterations and impaired sperm parameters due to carbamazepine usage.

Effects of intracerebroventricular administration of irisin on the hypothalamus-pituitary-gonadal axis in male rats.

Irisin is a product of fibronectin type III domain-containing protein 5 (FNDC5) and plays an important role in energy homeostasis. In this study, we aimed to determine effects of intracerebroventricular administration of irisin on the hypothalamus-pituitary-gonadal axis by molecular, biochemical, and morphological findings. Fourty male Wistar-Albino rats were used and divided into four groups including control, sham (vehicle), 10, and 100 nM irisin infused groups (n = 10). Hypothalamic gonadotropin releasing hormone (GnRH) level and serum luteinizing hormone (LH), follicle-stimulating hormone (FSH), and testosterone levels were determined. Testicular tissue histology and spermiogram analysis were also performed. Both irisin concentrations significantly reduced hypothalamic GnRH messenger RNA (mRNA) and protein levels (p < 0.05). It was found that serum LH, FSH, and testosterone levels and Sertoli and Leydig cell numbers were decreased by irisin administration (p < 0.05). In addition, irisin administration reduced sperm density and mobility (p < 0.05). However, it did not cause any change in testicular and epididymis weights and tubular diameter. Our results reveal that irisin can play a role in the central regulation of reproductive behavior and also reduces testosterone levels by suppressing LH and FSH secretion. These results suggest that the discovery of irisin receptor antagonists may be beneficial in the treatment of infertility.

Molecular circuit involving KLK4 integrates androgen and mTOR signaling in prostate cancer.

The androgen receptor (AR) and the phosphoinositide 3-kinase (PI3K)/protein kinase B/mammalian target of rapamycin (mTOR) signaling are two of the major proliferative pathways in a number of tissues and are the main therapeutic targets in various disorders, including prostate cancer (PCa). Previous work has shown that there is reciprocal feedback regulation of PI3K and AR signaling in PCa, suggesting that cotargeting both pathways may enhance therapeutic efficacy. Here we show that proteins encoded by two androgen-regulated genes, kallikrein related peptidase 4 (KLK4) and promyelocytic leukemia zinc finger (PLZF), integrate optimal functioning of AR and mTOR signaling in PCa cells. KLK4 interacts with PLZF and decreases its stability. PLZF in turn interacts with AR and inhibits its function as a transcription factor. PLZF also activates expression of regulated in development and DNA damage responses 1, an inhibitor of mTORC1. Thus, a unique molecular switch is generated that regulates both AR and PI3K signaling. Consistently, KLK4 knockdown results in a significant decline in PCa cell proliferation in vitro and in vivo, decreases anchorage-independent growth, induces apoptosis, and dramatically sensitizes PCa cells to apoptosis-inducing agents. Furthermore, in vivo nanoliposomal KLK4 siRNA delivery in mice bearing PCa tumors results in profound remission. These results demonstrate that the activities of AR and mTOR pathways are maintained by KLK4, which may thus be a viable target for therapy.

Paraneoplastic thrombocytosis in ovarian cancer.

BACKGROUND: The mechanisms of paraneoplastic thrombocytosis in ovarian cancer and the role that platelets play in abetting cancer growth are unclear. METHODS: We analyzed clinical data on 619 patients with epithelial ovarian cancer to test associations between platelet counts and disease outcome. Human samples and mouse models of epithelial ovarian cancer were used to explore the underlying mechanisms of paraneoplastic thrombocytosis. The effects of platelets on tumor growth and angiogenesis were ascertained. RESULTS: Thrombocytosis was significantly associated with advanced disease and shortened survival. Plasma levels of thrombopoietin and interleukin-6 were significantly elevated in patients who had thrombocytosis as compared with those who did not. In mouse models, increased hepatic thrombopoietin synthesis in response to tumor-derived interleukin-6 was an underlying mechanism of paraneoplastic thrombocytosis. Tumor-derived interleukin-6 and hepatic thrombopoietin were also linked to thrombocytosis in patients. Silencing thrombopoietin and interleukin-6 abrogated thrombocytosis in tumor-bearing mice. Anti-interleukin-6 antibody treatment significantly reduced platelet counts in tumor-bearing mice and in patients with epithelial ovarian cancer. In addition, neutralizing interleukin-6 significantly enhanced the therapeutic efficacy of paclitaxel in mouse models of epithelial ovarian cancer. The use of an antiplatelet antibody to halve platelet counts in tumor-bearing mice significantly reduced tumor growth and angiogenesis. CONCLUSIONS: These findings support the existence of a paracrine circuit wherein increased production of thrombopoietic cytokines in tumor and host tissue leads to paraneoplastic thrombocytosis, which fuels tumor growth. We speculate that countering paraneoplastic thrombocytosis either directly or indirectly by targeting these cytokines may have therapeutic potential. (Funded by the National Cancer Institute and others.).

Liraglutide Treatment in a Morbidly Obese Adolescent with a MC4R Gene Variant: Side Effects Reduce Success

Variants of the melanocortin-4 receptor (MC4R) gene are the most common cause of monogenic obesity. It has been shown that, while obesity cannot be controlled with diet and exercise, glucagon-like-peptide-1 receptor agonists (GLP-1 RA) provide weight loss in the short term. In this paper, our experience with liraglutide treatment in an adolescent patient carrying a MC4R gene variant is presented. A female patient was admitted first at the age of 12.5 years with a complaint of progressive weight gain. She had marked excess of appetite since infancy. On physical examination of the pubertal female patient with a body mass index (BMI) of 36.1 kg/m2 (3.48 standard deviation score), there was no pathological finding except diffuse acanthosis nigricans. Laboratory examinations revealed only insulin resistance. Weight loss was not achieved with lifestyle changes, metformin and orlistat treatments. On genetic examination, a sporadic heterozygous c.206T>G(p.I69R) variant that had been reported previously, was found in MC4R gene. Treatment with the GLP-1 RA, liraglutide, was initiated and a 19.2% reduction was achieved in the body weight and BMI at the end of 32 weeks. However, the patient, whose treatment compliance was disrupted due to significant gastrointestinal complaints, returned to her former weight within a few months (13 weeks) after treatment was stopped. In this case with a known pathogenic variant in MC4R gene, decrease of appetite and weight loss were achieved with liraglutide treatment, but side-effects of this treatment led to discontinuation of therapy. In such cases, there is need for effective and tolerable treatment options.

Single-center experience in management of progressive familial intrahepatic cholestasis.

BACKGROUND AND STUDY AIMS: Progressive familial intrahepatic cholestasis (PFIC) is an autosomal recessively inherited disease that causes intrahepatic-hepatocellular cholestasis. PFIC constitutes approximately 10-15% of cholestatic liver diseases in children. The aim of this study is to draw attention to this group of diseases, which pose a higher risk, in societies where consanguineous marriage is more common, and to share our experiences since the studies in the literature, regarding this group of diseases are case series with small number of patients. PATIENTS AND METHODS: This cross-sectional study was conducted on 34 patients who were admitted with jaundice and diagnosed by genetic analysis, between January 2015 and July 2020. RESULTS: We found 17.6% of patients with PFIC type 1, 55.9% patients had PFIC type 2, 14.7% patients had PFIC type 3, 8.8% patients had PFIC type 4 and 2.9% patients had PFIC type 5. Partial internal biliary diversion was performed in 5 (14.7%) patients, who had severe itching during follow-up, did not respond to medical treatment, and did not have significant fibrosis in liver biopsy yet. The degree of itching before PIBD was rated as +4 (cutaneous erosion, bleeding and scarring), in 5 patients and the rates were 0 (absent) in two patients, and +1 (mild itching) in 3 patients, 6 months after PIBD, these differences were statistically significant(p = 0.027). The mean weight z score was-1.43 (-3.72-+0.73), before PIBD, while it was 0.39(-1.86 -+2.45), six months after PIBD; the diference was statistically significant(p = 0.043). Liver transplantation was performed in 12 (35.3%) patients with significant fibrosis in liver biopsy and developing signs of portal hypertension. CONCLUSION: The PFIC disease group is a heterogeneous disease group that is difficult to diagnose and treat. It should be considered in patients with cholestasis and/or pruritus and those with a history of consanguineous marriage between parents and death of a sibling with similar clinical symptoms.

Liposomal 2-Methoxyestradiol Nanoparticles for Treatment of Uterine Leiomyoma in a Patient-Derived Xenograft Mouse Model.

Uterine leiomyomas represent a challenging problem with limited medical treatment options. The anti-tumor agent 2-methoxyestradiol (2-ME) shows promising results but its efficacy is limited by inadequate pharmacokinetics. We previously demonstrated that 2-ME nanoparticles can be successfully formulated and that they show improved in vitro anti-leiomyoma cell activity. Here, we examined the effects of the in vivo delivery of 2-ME nanoparticles in a patient-derived xenograft (PDX) leiomyoma mouse model. Patient-derived leiomyoma tumor tissues were xenografted subcutaneously in estrogen/progesterone pretreated immunodeficient NOG mice. Animals (n = 12) were treated with liposomal 2-ME nanoparticles by intra-peritoneal (IP) injection (50 mg/kg/dose, three times weekly) or control for 28 days. Tumor volume was measured weekly by calipers and prior to sacrifice by ultrasound. In addition, the expression of the cell proliferation marker Ki67 and the apoptosis marker cleaved caspase-3 in tumor tissues after treatment were measured by immunohistochemistry. Liposomal 2-ME treatment was associated with a significant tumor growth inhibition (30.5% less than controls as early as 2 weeks, p = 0.025). In addition, injections of liposomal 2-ME inhibited the expression of the proliferation marker Ki67 (55.8% reduction, p < 0.001). Furthermore, liposomal 2-ME treatment was associated with a 67.5% increase of cleaved caspase-3 expression of increase (p = 0.048). Our findings suggest that liposomal nanoparticle formulation can successfully deliver 2-ME and can be a promising therapeutic strategy for uterine leiomyoma. Further characterization of the liposomal-2ME, including pharmacokinetics, maximal tolerated dose, and safety, is needed in preclinical models prior to clinical trials.

A Case of Cleidocranial Dysplasia with a Novel Mutation and Growth Velocity Gain with Growth Hormone Treatment

Cleidocranial dysplasia (CCD) is a rare congenital autosomal dominant skeletal disorder that is characterized by hypoplasia or aplasia of clavicles, failure of cranial suture closure, dental anomalies, short stature and other changes in skeletal patterning and growth. The gene responsible for pathogenesis has been mapped to the short arm of chromosome 6p21, core binding factor alpha-1 (CBFA1) or runt related transcription factor-2 (RUNX2). Here we describe a CCD patient with a novel mutation in the RUNX2 gene. A five-and-a-half year old girl presented with severe short stature, dysmorphic facial appearance (hypertelorism, prominent forehead, high palate, midfacial hypoplasia), macrocephaly, large anterior fontanelle, increased anteroposterior chest diameter. Her shoulders were close to each other and her bilateral clavicles appeared short on physical examination. Bilateral hypoplastic clavicles, coxa valga, hypoplasia of iliac bones, wide symphysis pubis and phalangeal dysplastic features were detected on her skeletal X-ray examination. She was diagnosed as having CCD. Molecular analysis detected a novel heterozygous mutation 'NM_001024630.3p.T155P(c.463A>C)' in the RUNX2 gene. At age seven years and two months old, because of her severe short stature, growth hormone (GH) treatment was started and she responded well to GH therapy with no adverse effects. In conclusion, hypoplasia or aplasia of the clavicles, failure of cranial suture closure, dental anomalies and short stature should bring CCD to mind. We present a novel mutation in the RUNX2 gene for CCD. We obtained growth velocity gain with GH treatment in our patient.

Autophagy is Required to Regulate Mitochondria Renewal, Cell Attachment, and All-trans-Retinoic Acid-Induced Differentiation in NB4 Acute Promyelocytic Leukemia Cells.

All-trans-retinoic acid (ATRA) is a potent inducer of cellular differentiation, growth arrest, and apoptosis as well as a front-line therapy for acute promyelocytic leukemia (APL). The present study provides evidence that induction of autophagy is required for ATRA to induce differentiation of APL (NB4) cells into granulocytes. ATRA treatment causes ~12-fold increase in the number of acidic vesicular organelles and induces marked up-regulation of LC3-II, autophagy-related 5 (ATG5), and Beclin-1. Transmission electron microscopy (TEM) revealed a decrease in mitochondria and ATRA-induced differentiation. To determine the role of autophagy in the differentiation of APL, we knocked down ATG5 in NB4 cells to find that ATRA-induced differentiation is significantly inhibited during ATG5 knock down in cells, indicating the role of autophagy in differentiation of APL. Further experiments revealed restriction of autophagy during ATRA-induced differentiation and inhibition of tissue transglutaminase 2 (TG2) and phospho-focal adhesion kinase (p-FAK), which are known to have roles in differentiation and cell attachment. We examined expression of Beclin-1 and B-cell lymphoma-2 (Bcl-2) and levels of mechanistic target of rapamycin (mTOR) after ATRA treatment. ATRA inhibits Bcl-2, up-regulates Beclin-1 expression, and reduces induction of mTOR activation/phosphorylation in NB4 cells. Our results reveal that autophagy has roles in regulation of differentiation, mitochondria elimination, and cell attachment during ATRA-induced APL differentiation.

Autosomal recessive cutis laxa: a novel mutation in the FBLN5 gene in a family.

FBLN5-related cutis laxa (CL) is a rare syndrome that can be inherited in an autosomal dominant or recessive manner. Autosomal recessive cutis laxa (ARCL), type IA, has been reported to be more severe. The disease is characterized by microcephaly, sagging cheeks, loose, wrinkled and redundant skin, emphysema, aorta or pulmonary artery abnormalities, inguinal hernia, and anomalies of internal organs. Homozygous mutations in the FBLN5 gene are responsible for the clinical manifestations. We report a family study of a child with ARCL. FBLN5 genes of the patient and parents were sequenced using next-generation sequencing technologies. Analyses showed that the patient was homozygous for the novel c.518A>G, p.R173H mutation in exon 6 of the FBLN5 gene, whereas the parents were heterozygous. The mutation was found to be 'possibly pathogenic' in bioinformatic analysis. We identified a novel FBLN5 mutation in a CL patient; pedigree and parental genetic analyses suggested ARCL. Our results also suggest that the mutation analysis provides useful evidence to support the clinical diagnosis and define the inheritance mode of CL in an apparently sporadic case.

Novel Mutations in Obesity-related Genes in Turkish Children with Non-syndromic Early Onset Severe Obesity: A Multicentre Study

Objective: Non syndromic monogenic obesity is a rare cause of early onset severe obesity in the childhood period. This form may not be distinguishable from other forms of severe obesity without genetic analysis, particularly if patients do not exibit any physical abnormalities or developmental delay. The aim of this study was to screen 41 different obesity-related genes in children with non-syndromic early onset severe obesity. Methods: Children with severe (body mass index-standard deviation score >3) and early onset (<7 years) obesity were screened by next-generation sequencing based, targeted DNA custom panel for 41 known-obesity-related genes and the results were confirmed by Sanger technique. Results: Six novel variants were identified in five candidate genes in seven out of 105 children with severe obesity; two in SIM1 (p.W306C and p.Q36X), one in POMC (p.Y160H), one in PCSK1 (p.W130G fs Ter8), two in MC4R (p.D126E) and one in LEPR (p.Q4H). Additionally, two previously known variations in MC4R were identified in four patients (p.R165W in three, and p.V166I in one). Conclusion: We identified six novel and four previously described variants in six obesity-related genes in 11 out of 105 childrens with early onset severe obesity. The prevalence of monogenic obesity was 10.4% in our cohort.

A mitochondrial neurogastrointestinal encephalomyopathy with intestinal pseudo-obstruction resulted from a novel splice site mutation.

Mitochondrial neurogastrointestinal encephalopathy (MNGIE) is an autosomal recessive disorder characterized by gastrointestinal dysmotility, cachexia, ptosis, peripheral neuropathy and leukoencephalopathy. The diagnosis is often not made until 5-10 years after the onset of symptoms. MNGIE is caused by mutations in thymidine phosphorylase gene TYMP. Here, we present a 19-year-old boy with MNGIE who had a chronic intestinal pseudo-obstruction, and we describe his family history. Genetic analysis revealed a novel homozygous c.765+1G>C intronic mutation which is expected to disrupt splicing of TYMP in the patient. Family screening revealed that the brother was also affected and the mother was a carrier. MNGIE should be considered and genetic testing instigated if individuals with cachexia have neuromuscular complaints or symptoms of chronic intestinal pseudo-obstruction.

Central irisin administration suppresses thyroid hormone production but increases energy consumption in rats.

Irisin, which is secreted from the skeletal muscle in response to physical exercise and defined as a thermogenic peptide, may play an important role in energy metabolism. Thyroid hormones, which are one of the other influential factors on the metabolic status, increase heat production and are the main regulators of energy metabolism. This study was conducted to determine the possible effects of irisin administration on thyroid hormones. Forty adult male Wistar albino rats were used in the study. The rats were equally divided into 4 groups (n = 10). The brain infusion kit was implanted in the groups, and irisin (or solvent as control) was centrally administered to the rats via osmotic mini pumps for 7 days. During the experiment, food consumption, body weights, and body temperatures of the animals were recorded. Food intake was significantly increased in the groups treated with irisin (p < 0.05), but their body weights were not changed. Hypothalamic TRH gene expression, serum TSH, fT3, and fT4 levels were significantly lower in the groups treated with irisin as compared to the naive and control groups (p < 0.05). In addition, irisin increased UCP1 mRNA expression in white and brown adipose tissue and UCP3 mRNA expression in muscle tissue in rats and also raised their body temperature (p < 0.05). Consequently, although central irisin administration has inhibitory effects on the hypothalamic-pituitary-thyroid axis, it seems to be an important agent in the regulation of food intake and energy metabolism.

Lack of association between pseudoexfoliation syndrome and manganese superoxide dismutase polymorphism.

PURPOSE: The aim of this clinical study was to investigate the possible association between manganese superoxide dismutase (Mn-SOD) enzyme polymorphism in the mitochondrial targeting sequence and pseudoexfoliation syndrome. METHODS: Ala (GTT) or Val (GCT) polymorphism in the signal peptide of Mn-SOD gene was evaluated using a primer pair to amplify a 107-bp fragment followed by digestion with restriction enzyme NgoM IV. RESULTS: The frequencies of Ala-9 and Val-9 variants and genotypes of Mn-SOD were similar in the controls and pseudoexfoliation syndrome patients. CONCLUSIONS: These results suggest no major modifying role for the Mn-SOD gene polymorphism in patients with pseudoexfoliation syndrome.

STAMP2 increases oxidative stress and is critical for prostate cancer.

The six transmembrane protein of prostate 2 (STAMP2) is an androgen-regulated gene whose mRNA expression is increased in prostate cancer (PCa). Here, we show that STAMP2 protein expression is increased in human PCa compared with benign prostate that is also correlated with tumor grade and treatment response. We also show that STAMP2 significantly increased reactive oxygen species (ROS) in PCa cells through its iron reductase activity which also depleted NADPH levels. Knockdown of STAMP2 expression in PCa cells inhibited proliferation, colony formation, and anchorage-independent growth, and significantly increased apoptosis. Furthermore, STAMP2 effects were, at least in part, mediated by activating transcription factor 4 (ATF4), whose expression is regulated by ROS. Consistent with in vitro findings, silencing STAMP2 significantly inhibited PCa xenograft growth in mice. Finally, therapeutic silencing of STAMP2 by systemically administered nanoliposomal siRNA profoundly inhibited tumor growth in two established preclinical PCa models in mice. These data suggest that STAMP2 is required for PCa progression and thus may serve as a novel therapeutic target.

The Ala-9Val polymorphism in the mitochondrial targeting sequence (MTS) of the manganese superoxide dismutase gene is not associated with juvenile-onset asthma.

BACKGROUND: We aimed to investigate the possible association between Mn-SOD polymorphism in the mitochondrial targeting sequence and asthma. METHODS: Alanine or valine polymorphism in the signal peptide of Mn-SOD gene was evaluated using a primer pair to amplify a 107-bp fragment followed by digestion with NgoM IV. RESULTS: No significant difference in genotype frequencies was found between patients and controls. CONCLUSION: These results suggest no major modifying role for the Mn-SOD gene polymorphism in patients with asthma.

Antagonism of tumoral prolactin receptor promotes autophagy-related cell death.

Therapeutic upregulation of macroautophagy in cancer cells provides an alternative mechanism for cell death. Prolactin (PRL) and its receptor (PRLR) are considered attractive therapeutic targets because of their roles as growth factors in tumor growth and progression. We utilized G129R, an antagonist peptide of PRL, to block activity of the tumoral PRL/PRLR axis, which resulted in inhibition of tumor growth in orthotopic models of human ovarian cancer. Prolonged treatment with G129R induced the accumulation of redundant autolysosomes in 3D cancer spheroids, leading to a type II programmed cell death. This inducible autophagy was a noncanonical beclin-1-independent pathway and was sustained by an astrocytic phosphoprotein (PEA-15) and protein kinase C zeta interactome. Lower levels of tumoral PRL/PRLR in clinical samples were associated with longer patient survival. Our findings provide an understanding of the mechanisms of tumor growth inhibition through targeting PRL/PRLR and may have clinical implications.