SS31 Ameliorates Oxidative Stress via the Restoration of Autophagic Flux to Protect Aged Mice From Hind Limb Ischemia.

Background: Oxidative stress and impaired autophagic flux play important roles in the development of peripheral artery disease (PAD). SS31 is considered an important antioxidant peptide and autophagy regulator. We aimed to investigate the role of SS31 in PAD myopathy and its possible mechanism both in vivo and in vitro. Methods: A hind limb ischemia (HLI) model was established with old C57BL/6 (14-month-old) mice. Mice in the SS31 group were intraperitoneally injected with SS31 (3 mg/kg) for 4 weeks. We examined skeletal muscle function and histomorphology, autophagy-related protein levels and reactive oxygen species (ROS) content. For the in vitro experiments, after C2C12 myotubes were treated with CoCl2, SS31, and chloroquine (CQ) or rapamycin (RAPA), we measured ROS content, autophagy-related protein levels and antioxidant enzyme expression. Results: SS31 treatment effectively enhanced the recovery of skeletal muscle function, alleviated skeletal muscle injury and suppressed mitochondrial ROS production in ischemic limbs. SS31 reduced apoptosis and oxidative stress, and SS31 restored impaired autophagic flux by inhibiting the AKT-mTOR pathway. In vitro studies showed that SS31 restored autophagic flux and improved oxidative stress in C2C12 cells. Moreover, phosphorylated AKT (p-AKT) and phosphorylated mTOR (p-mTOR) levels were reduced. Conclusion: These experiments indicated that SS31 can inhibit oxidative stress by restoring autophagic flux to reverse hypoxia-induced injury in vivo and in vitro.

Machine learning-based statistical analysis for early stage detection of cervical cancer.

Cervical cancer (CC) is the most common type of cancer in women and remains a significant cause of mortality, particularly in less developed countries, although it can be effectively treated if detected at an early stage. This study aimed to find efficient machine-learning-based classifying models to detect early stage CC using clinical data. We obtained a Kaggle data repository CC dataset which contained four classes of attributes including biopsy, cytology, Hinselmann, and Schiller. This dataset was split into four categories based on these class attributes. Three feature transformation methods, including log, sine function, and Z-score were applied to these datasets. Several supervised machine learning algorithms were assessed for their performance in classification. A Random Tree (RT) algorithm provided the best classification accuracy for the biopsy (98.33%) and cytology (98.65%) data, whereas Random Forest (RF) and Instance-Based K-nearest neighbor (IBk) provided the best performance for Hinselmann (99.16%), and Schiller (98.58%) respectively. Among the feature transformation methods, logarithmic gave the best performance for biopsy datasets whereas sine function was superior for cytology. Both logarithmic and sine functions performed the best for the Hinselmann dataset, while Z-score was best for the Schiller dataset. Various Feature Selection Techniques (FST) methods were applied to the transformed datasets to identify and prioritize important risk factors. The outcomes of this study indicate that appropriate system design and tuning, machine learning methods and classification are able to detect CC accurately and efficiently in its early stages using clinical data.

100 years of iodine testing of the cervix: A critical review and implications for the future.

OBJECTIVES: We aim to describe the history of iodine testing of the cervix and identify areas where further work is required. STUDY DESIGN: We conducted a search of PubMed and Google Scholar. Full article texts were reviewed. Reference lists were screened for additional articles and books. 37 basic articles in journals including ones written in German and three basic articles in books were identified. RESULTS: Glycogen staining of the ectocervical squamous epithelium with iodine goes back to Paul Ehrlich (1854-1915). Walter Schiller (1887-1960) examined nearly 200 different dyes and found that vital staining of the cervical squamous epithelium was best achieved with Lugol's iodine solution, which was indicated by Jean Guillaume Lugol (1786-1851) for disinfection of the vagina. In 1928 W. Lahm observed that the glycogen content of a squamous epithelium cell decreases as anaplasia increases. From the outset, H. Hinselmann included the iodine test in the minimum requirements for colposcopy. In 1946 H. J. Wespi first mentioned the finding of an "uncharacteristic iodine negative area." The first international colposcopic terminology from Graz in 1975 lists the "iodine light area" among the different colposcopy findings. The IFCPC nomenclatures from Rome 1990, Barcelona 2002, and Rio de Janeiro 2011 have evaluated the iodine test and classified their findings differently. A breakthrough to effective cervical cancer screening in resource-limited settings in Africa, India, and Latin America was achieved with R. Sankaranarayanan's publication on naked-eye visual inspection of the cervix after application of Lugol's iodine. CONCLUSIONS: This paper is a step toward a better understanding of what we think and do today with iodine testing and what problems and upcoming tasks will arise in future.

Peptide Szeto­Schiller 31 ameliorates doxorubicin­induced cardiotoxicity by inhibiting the activation of the p38 MAPK signaling pathway.

Oxidative stress serves a key role in doxorubicin (DOX)­induced cardiotoxicity. The peptide Szeto­Schiller (SS)31 is an efficacious antioxidant with the capacity to reduce mitochondrial reactive oxygen species (ROS) levels and scavenge free radicals. Although SS31 is involved in the pathophysiological process of various cardiovascular diseases, the role of SS31 in DOX­induced cardiotoxicity remains unclear. To explore the effects of SS31 in DOX­induced cardiotoxicity, the present study first constructed DOX­induced cardiotoxicity models, in which H9c2 cells were incubated with 1 µM DOX for 24 h and C57BL/6 mice were administered DOX (20 mg/kg cumulative dose). The results of various assays in these models demonstrated that SS31 exhibited a cardioprotective effect in vitro and in vivo by attenuating the level of ROS, stabilizing the mitochondrial membrane potential and ameliorating myocardial apoptosis as well as fibrosis following treatment with DOX. Mechanistically, the results of the present study revealed that the p38 MAPK signaling pathway was inhibited by SS31 in DOX­treated H9c2 cells, which was associated with the cardioprotective function of SS31. In addition, P79350, a selective agonist of p38 MAPK, reversed the protective effects of SS31. Taken together, these results demonstrated the effects of SS31 on ameliorating DOX­induced cardiotoxicity and indicated its potential as a drug for the treatment of DOX­induced cardiotoxicity.

The Mitochondrion-Targeted Antioxidants in Kidney Disease.

Mitochondria are a potent source of cellular reactive oxygen species (ROS) and are vulnerable to oxidative damage. Mitochondria dysfunction could result in adenosine triphosphate (ATP) decrease and cell death. The kidney is an ATP-consuming organ, and the relationship between mitochondrial dysfunction and renal disease has been long noted. Mitochondrial targeting is a novel strategy for kidney diseases. At present, there are several ways to target mitochondria, such as the addition of a triphenylphosphonium cation, mitochondria-targeted peptides, and nanocarrier. There are also a variety of choices for the payload, such as nitroxides, quinone derivates, vitamins and so on. This review summarized the chemical and also clinical characteristics of various mitochondria- targeted antioxidants and focused on their application and perspectives in kidney diseases.

The mitochondria-targeted peptide SS-31 binds lipid bilayers and modulates surface electrostatics as a key component of its mechanism of action.

Mitochondrial dysfunction underlies many heritable diseases, acquired pathologies, and aging-related declines in health. Szeto-Schiller (SS) peptides comprise a class of amphipathic tetrapeptides that are efficacious toward a wide array of mitochondrial disorders and are believed to target mitochondrial membranes because they are enriched in the anionic phospholipid cardiolipin (CL). However, little is known regarding how SS peptides interact with or alter the physical properties of lipid bilayers. In this study, using biophysical and computational approaches, we have analyzed the interactions of the lead compound SS-31 (elamipretide) with model and mitochondrial membranes. Our results show that this polybasic peptide partitions into the membrane interfacial region with an affinity and a lipid binding density that are directly related to surface charge. We found that SS-31 binding does not destabilize lamellar bilayers even at the highest binding concentrations; however, it did cause saturable alterations in lipid packing. Most notably, SS-31 modulated the surface electrostatics of both model and mitochondrial membranes. We propose nonexclusive mechanisms by which the tuning of surface charge could underpin the mitoprotective properties of SS-31, including alteration of the distribution of ions and basic proteins at the interface, and/or modulation of bilayer physical properties. As a proof of concept, we show that SS-31 alters divalent cation (calcium) distribution within the interfacial region and reduces the energetic burden of calcium stress in mitochondria. The mechanistic details of SS-31 revealed in this study will help inform the development of future compound variants with enhanced efficacy and bioavailability.

Educational Case: Yolk Sac (Endodermal Sinus) Tumor of the Ovary.

The following fictional case is intended as a learning tool within the Pathology Competencies for Medical Education (PCME), a set of national standards for teaching pathology. These are divided into three basic competencies: Disease Mechanisms and Processes, Organ System Pathology, and Diagnostic Medicine and Therapeutic Pathology. For additional information, and a full list of learning objectives for all three competencies, see http://journals.sagepub.com/doi/10.1177/2374289517715040.1.

Gastric yolk sac tumor with synchronous liver metastasis: first case report in China.

Gastric yolk sac tumors (YSTs) are extremely rare and often present a grave clinical course at the time of diagnosis. The prognosis of gastric YSTs is poor; the effective therapeutic approach beyond early diagnosis and curative resection is still uncertain. Here we first report a case of gastric YST without relevant clinical symptoms, presenting liver mass as the first performance. The patient was diagnosed with gastric YST with synchronous liver metastasis and underwent simultaneous resection of the primary gastric lesion and liver metastasis successfully. To the best of our knowledge, this is the first case report of gastric YST in China. AFP is a prognostic marker for recurrence and survival of gastric YST. Histologically, a reticular pattern (a clear-cell endoblastic pattern) with glomerulus-like structures (Schiller-Duval bodies) is golden criteria of diagnosis for YST. Our case will contribute to understanding the clinical characteristics of this rare disease.

Mitochondria-targeting drug conjugates for cytotoxic, anti-oxidizing and sensing purposes: current strategies and future perspectives.

Mitochondrial targeting is a promising approach for solving current issues in clinical application of chemotherapy and diagnosis of several disorders. Here, we discuss direct conjugation of mitochondrial-targeting moieties to anticancer drugs, antioxidants and sensor molecules. Among them, the most widely applied mitochondrial targeting moiety is triphenylphosphonium (TPP), which is a delocalized cationic lipid that readily accumulates and penetrates through the mitochondrial membrane due to the highly negative mitochondrial membrane potential. Other moieties, including short peptides, dequalinium, guanidine, rhodamine, and F16, are also known to be promising mitochondrial targeting agents. Direct conjugation of mitochondrial targeting moieties to anticancer drugs, antioxidants and sensors results in increased cytotoxicity, anti-oxidizing activity and sensing activity, respectively, compared with their non-targeting counterparts, especially in drug-resistant cells. Although many mitochondria-targeted anticancer drug conjugates have been investigated in vitro and in vivo, further clinical studies are still needed. On the other hand, several mitochondria-targeting antioxidants have been analyzed in clinical phases I, II and III trials, and one conjugate has been approved for treating eye disease in Russia. There are numerous ongoing studies of mitochondria-targeted sensors.

Cardiolipin-targeted peptides rejuvenate mitochondrial function, remodel mitochondria, and promote tissue regeneration during aging.

It has been proposed that a loss of bioenergetic capacity of cells contributes to the progressive loss of biological function with age. Aging is associated with loss of mitochondrial cristae membranes and inhibition of ATP production. Despite the many approaches being pursued for improving mitochondrial function, none of them directly targets the electron transport chain to improve ATP production. Recent studies have brought attention to cardiolipin as a unique target for promoting mitochondrial efficiency. Cardiolipin is important for cristae curvatures and is necessary for optimal activity of the respiratory complexes and the assembly of supercomplexes. Here we describe the discovery of a class of cell-penetrating aromatic-cationic tetrapeptides that selectively target cardiolipin and increase coupling efficiency while reducing reactive oxygen species production. These compounds can rejuvenate mitochondrial bioenergetics, remodel mitochondrial cristae structure, repair cellular structure, and restore organ function during aging.

Mitochondrial regulation of diabetic vascular disease: an emerging opportunity.

Diabetes-related vascular complication rates remain unacceptably high despite guideline-based medical therapies that are significantly more effective in individuals without diabetes. This critical gap represents an opportunity for researchers and clinicians to collaborate on targeting mechanisms and pathways that specifically contribute to vascular pathology in patients with diabetes mellitus. Dysfunctional mitochondria producing excessive mitochondrial reactive oxygen species (mtROS) play a proximal cell-signaling role in the development of vascular endothelial dysfunction in the setting of diabetes. Targeting the mechanisms of production of mtROS or mtROS themselves represents an attractive method to reduce the prevalence and severity of diabetic vascular disease. This review focuses on the role of mitochondria in the development of diabetic vascular disease and current developments in methods to improve mitochondrial health to improve vascular outcomes in patients with DM.

Mitochondrial bioenergetics, redox state, dynamics and turnover alterations in renal mass reduction models of chronic kidney diseases and their possible implications in the progression of this illness.

Nowadays, chronic kidney disease (CKD) is considered a worldwide public health problem. CKD is a term used to describe a set of pathologies that structurally and functionally affect the kidney, it is mostly characterized by the progressive loss of kidney function. Current therapeutic approaches are insufficient to avoid the development of this disease, which highlights the necessity of developing new strategies to reverse or at least delay CKD progression. Kidney is highly dependent on mitochondrial homeostasis and function, consequently, the idea that mitochondrial pathologies could play a pivotal role in the genesis and development of kidney diseases has risen. Although many research groups have recently published studies of mitochondrial function in acute kidney disease models, the existing information about CKD is still limited, especially in renal mass reduction (RMR) models. This paper focuses on reviewing current experimental information about the bioenergetics, dynamics (fission and fusion processes), turnover (mitophagy and biogenesis) and redox mitochondrial alterations in RMR, to discuss and integrate the mitochondrial changes triggered by nephron loss, as well as its relationship with loss of kidney function in CKD, in these models. Understanding these mechanisms would allow us to design new therapies that target these mitochondrial alterations.

Detection of precancerous lesions in the cervix and HPV infection in women in the region of Maniapure, Bolivar State.

Human papillomavirus (HPV) is the causative agent of cervical cancer (CC), the second most common cause of cancer deaths in Venezuela. Early detection and prompt treatment of precancerous lesions prevent up to 80% of CC cases. In Venezuela, difficult access to CC screening means that the disease is detected at advanced stages, especially in more vulnerable indigenous populations. The aim of the study was to detect precancerous cervical lesions and HPV infection in 60 women who attended the gynaecology service at the Maniapure Outpatient Clinic in Bolivar State, Venezuela. The study was carried out to detect precancerous cervical lesions using visual inspection with acetic acid (VIA), the Schiller test and conventional cytology (Pap testing). HPV detection and typing were carried out using the polymerase chain reaction. 58.3% of the women in the study belonged to the Eñepa indigenous community and 41.7% were white Creole women. The Schiller test showed irregularities in the staining of the exocervical epithelium in 8.33% of the patients, suggesting HPV infection. VIA was positive for 10.0% of the women. In the cytopathology report, 81.67% tested negative for intraepithelial lesions. The overall frequency of HPV detection was 35.0%. HPV infection was detected in 45.71% of the Eñepa women and 20.0% of the Creole women. 71.43% of the women had a high-risk single HPV infection. The percentage of viral infection was lower in the Creole patients than in the indigenous population; therefore, CC screening programmes in the latter population need to be improved.

Tumor germinal ovárico tipo seno endodérmico en adolescente / Ovarian germ cell tumor of the endodermal sinus type in adolescents

Las afecciones tumorales de ovario en niñas son infrecuentes. El tumor del seno endodérmico es un tumor derivado de células multipotenciales primitivas o células germinales, que se diferencian en estructuras del saco vitelino. Es muy agresivo y poco frecuente. Se presenta el caso de una paciente femenina, de 15 años de edad, delgada, con marcada palidez cutáneo mucosa, masa abdominal palpable, que ocupa todo el hemiabdomen inferior desde hipogastrio hasta tres cm por encima del ombligo, no dolorosa a la palpación, firme y superficie lisa con borde irregular y poco movible. Se indicaron estudios complementarios e imagenológicos. La tomografía axial computarizada informó gran masa tumoral intraabdominal. Se realizó excéresis del tumor y estudio anatomopatológico que concluyó diagnóstico de tumor del seno endodérmico o del saco vitelino del ovario derecho que se acompaña de necrosis y extensas zonas de hemorragias. Se presenta así a la comunidad médica, como el primer caso con diagnóstico de esta entidad en la provincia(AU)

Ovarian tumor diseases in girls are infrequent. Endodermal sinus tumor is a tumor derived from primitive multipotential cells or germ cells, which differentiate into structures of the yolk sac. The tumor is very aggressive and rare. This study presents the case of a 15-year-old female patient with a chief complaint of increase of the lower abdomen and moderate abdominal pain in the epigastrium. She was thin, pale and had a palpable abdominal mass in the lower hemiabdomen from the hypogastrium up to 3 cm above the navel. On palpation the mass was not painful, and it was firm and smooth with irregular border and unmovable. Investigations were ordered, including imaging tests. Surgical intervention was decided to perform excision of the tumor and anatomopathologic studies. A diagnosis of endodermal sinus tumor or yolk sac tumor of the right ovary was made. This study is presented to the medical community as the first case with a diagnosis of this condition in the province(AU)

Pure Primary Extragonadal Retroperitoneal Yolk Sac Tumour in a Young Child: A Case Report.

Germ Cell Tumours (GCTs) in children are uncommon, constituting approximately only 3% of all malignancies in children younger than 15 years of age. Primary extragonadal GCTs constitute only 1-5% of all GCTs and a retroperitoneal site is seen only in 4% of all extragonadal GCTs. Extragonadal GCTs arise from local transformation of primordial germ cells which have been misplaced during the migration of these cells through the midline dorsal mesentery in the fourth-sixth week of embryogenesis. GCTs in children show remarkable variability in age, site, presentation and histology. This is the case of a three-year-old male child who presented with a history of an abdominal swelling which was rapidly progressive in nature. Radiology showed a large retroperitoneal mass and lesions in the liver. Histopathology, immunohistochemistry and serum Alpha-fetoprotein (AFP) values confirmed a diagnosis of pure primary extragonadal yolk sac tumour.

Primary Nasopharngeal Yolk Sac Tumor: A Case Report.

Yolk sac tumour also known as primitive endodermal tumour is the most common malignant germ cell tumour (GCT) in the paediatric age group. Most common sites of involvement are ovaries and testes, but rarely can occur in the extragonadal sites. In the head and neck region, yolk sac tumours have been reported in the nasopharynx, sinonasal tract, orbit, ear and parotid gland. Nasopharynx is an uncommon site for yolk sac tumour and very few cases of nasopharngeal pure yolk sac tumour have been reported so far. Yolk sac tumours are highly malignant and have a poor prognosis. This is a case of pure GCT in a three-year-old female child who presented with a rapidly growing nasopharyngeal mass. Histopathological examination followed by immunohistochemistry and serum AFP study clinched the diagnosis of yolk sac tumour. The tumour responded well to chemotherapy as evidenced by decrease in serum AFP levels.

Improving mitochondrial bioenergetics under ischemic conditions increases warm ischemia tolerance in the kidney.

Ischemia time during partial nephrectomy is strongly associated with acute and chronic renal injury. ATP depletion during warm ischemia inhibits ATP-dependent processes, resulting in cell swelling, cytoskeletal breakdown, and cell death. The duration of ischemia tolerated by the kidney depends on the amount of ATP that can be produced with residual substrates and oxygen in the tissue to sustain cell function. We previously reported that the rat can tolerate 30-min ischemia quite well but 45-min ischemia results in acute kidney injury and progressive interstitial fibrosis. Here, we report that pretreatment with SS-20 30 min before warm ischemia in the rat increased ischemia tolerance from 30 to 45 min. Histological examination of kidney tissues revealed that SS-20 reduced cytoskeletal breakdown and cell swelling after 45-min ischemia. Electron microscopy showed that SS-20 reduced mitochondrial matrix swelling and preserved cristae membranes, suggesting that SS-20 enhanced mitochondrial ATP synthesis under ischemic conditions. Studies with isolated kidney mitochondria showed dramatic reduction in state 3 respiration and respiratory control ratio after 45-min ischemia, and this was significantly improved by SS-20 treatment. These results suggest that SS-20 increases efficiency of the electron transport chain and improves coupling of oxidative phosphorylation. SS-20 treatment after ischemia also significantly reduced interstitial fibrosis. These new findings reveal that enhancing mitochondrial bioenergetics may be an important target for improving ischemia tolerance, and SS-20 may serve well for minimizing acute kidney injury and chronic kidney disease following surgical procedures such as partial nephrectomy and transplantation.

Novel cardiolipin therapeutic protects endothelial mitochondria during renal ischemia and mitigates microvascular rarefaction, inflammation, and fibrosis.

Microvascular rarefaction, or loss of microvascular density, is increasingly implicated in the progression from acute ischemic kidney injury to chronic kidney disease. Microvascular dropout results in chronic tissue hypoxia, interstitial inflammation, and fibrosis. There is currently no therapeutic intervention for microvascular rarefaction. We hypothesize that capillary dropout begins with ischemic damage to endothelial mitochondria due to cardiolipin peroxidation, resulting in loss of cristae and the failure to regenerate ATP upon reperfusion. SS-31 is a cell-permeable peptide that targets the inner mitochondrial membrane and binds selectively to cardiolipin. It was recently shown to inhibit cardiolipin peroxidation by cytochrome c peroxidase activity, and it has been shown to protect mitochondrial cristae in proximal tubular cells during ischemia, and accelerated ATP recovery upon reperfusion. We found mitochondrial swelling and loss of cristae membranes in endothelial and medullary tubular epithelial cells after 45-min ischemia in the rat. The loss of cristae membranes limited the ability of these cells to regenerate ATP upon reperfusion and led to loss of vascular integrity and to tubular cell swelling. SS-31 prevented mitochondria swelling and protected cristae membranes in both endothelial and epithelial cells. By minimizing endothelial and epithelial cell injury, SS-31 prevented "no-reflow" after ischemia and significantly reduced the loss of peritubular capillaries and cortical arterioles, interstitial inflammation, and fibrosis at 4 wk after ischemia. These results suggest that mitochondria protection represents an upstream target for pharmacological intervention in microvascular rarefaction and fibrosis.

Targeting mitochondrial cardiolipin and the cytochrome c/cardiolipin complex to promote electron transport and optimize mitochondrial ATP synthesis.

BACKGROUND AND PURPOSE: Cardiolipin plays an important role in mitochondrial respiration and cardiolipin peroxidation is associated with age-related diseases. Hydrophobic interactions between cytochrome c and cardiolipin converts cytochrome c from an electron carrier to a peroxidase. In addition to cardiolipin peroxidation, this impedes electron flux and inhibits mitochondrial ATP synthesis. SS-31 (D-Arg-dimethylTyr-Lys-Phe-NH2 ) selectively binds to cardiolipin and inhibits cytochrome c peroxidase activity. Here, we examined whether SS-31 also protected the electron carrier function of cytochrome c. EXPERIMENTAL APPROACH: Interactions of SS-31 with cardiolipin were studied using liposomes and bicelles containing phosphatidylcholine alone or with cardiolipin. Structural interactions were assessed by fluorescence spectroscopy, turbidity and nuclear magnetic resonance. Effects of cardiolipin on electron transfer kinetics of cytochrome c were determined by cytochrome c reduction in vitro and oxygen consumption using mitoplasts, frozen and fresh mitochondria. KEY RESULTS: SS-31 interacted only with liposomes and bicelles containing cardiolipin in about 1:1 ratio. NMR studies demonstrated that the aromatic residues of SS-31 penetrated deep into cardiolipin-containing bilayers. SS-31 restored cytochrome c reduction and mitochondrial oxygen consumption in the presence of added cardiolipin. In fresh mitochondria, SS-31 increased state 3 respiration and efficiency of ATP synthesis. CONCLUSIONS AND IMPLICATIONS: SS-31 selectively targeted cardiolipin and modulated its interaction with cytochrome c. SS-31 inhibited the cytochrome c/cardiolipin complex peroxidase activity while protecting its ability to serve as an electron carrier, thus optimizing mitochondrial electron transport and ATP synthesis. This novel class of cardiolipin therapeutics has the potential to restore mitochondrial bioenergetics for treatment of numerous age-related diseases.