Comparison of the transcriptome, lipidome, and c-di-GMP production between BCGΔBCG1419c and BCG, with Mincle- and Myd88-dependent induction of proinflammatory cytokines in murine macrophages.

We have previously reported the transcriptomic and lipidomic profile of the first-generation, hygromycin-resistant (HygR) version of the BCGΔBCG1419c vaccine candidate, under biofilm conditions. We recently constructed and characterized the efficacy, safety, whole genome sequence, and proteomic profile of a second-generation version of BCGΔBCG1419c, a strain lacking the BCG1419c gene and devoid of antibiotic markers. Here, we compared the antibiotic-less BCGΔBCG1419c with BCG. We assessed their colonial and ultrastructural morphology, biofilm, c-di-GMP production in vitro, as well as their transcriptomic and lipidomic profiles, including their capacity to activate macrophages via Mincle and Myd88. Our results show that BCGΔBCG1419c colonial and ultrastructural morphology, c-di-GMP, and biofilm production differed from parental BCG, whereas we found no significant changes in its lipidomic profile either in biofilm or planktonic growth conditions. Transcriptomic profiling suggests changes in BCGΔBCG1419c cell wall and showed reduced transcription of some members of the DosR, MtrA, and ArgR regulons. Finally, induction of TNF-α, IL-6 or G-CSF by bone-marrow derived macrophages infected with either BCGΔBCG1419c or BCG required Mincle and Myd88. Our results confirm that some differences already found to occur in HygR BCGΔBCG1419c compared with BCG are maintained in the antibiotic-less version of this vaccine candidate except changes in production of PDIM. Comparison with previous characterizations conducted by OMICs show that some differences observed in BCGΔBCG1419c compared with BCG are maintained whereas others are dependent on the growth condition employed to culture them.

Transient inhibition of sodium-glucose cotransporter 2 after ischemia/reperfusion injury ameliorates chronic kidney disease.

Sodium-glucose cotransporter 2 (SGLT2) inhibitor, dapagliflozin (Dapa), exhibited nephroprotective effects in patients with chronic kidney disease (CKD). We assessed the efficacy of short-term Dapa administration following acute kidney injury (AKI) in preventing CKD. Male Wistar rats were randomly assigned to Sham surgery, bilateral ischemia for 30 minutes (abbreviated as IR), and IR + Dapa groups. Daily treatment with Dapa was initiated just 24 hours after IR and maintained for only 10 days. Initially, rats were euthanized at this point to study early renal repair. After severe AKI, Dapa promptly restored creatinine clearance (CrCl) and significantly reduced renal vascular resistance compared with the IR group. Furthermore, Dapa effectively reversed the mitochondrial abnormalities, including increased fission, altered mitophagy, metabolic dysfunction, and proapoptotic signaling. To study this earlier, another set of rats was studied just 5 days after AKI. Despite persistent renal dysfunction, our data reveal a degree of mitochondrial protection. Remarkably, a 10-day treatment with Dapa demonstrated effectiveness in preventing CKD transition in an independent cohort monitored for 5 months after AKI. This was evidenced by improvements in proteinuria, CrCl, glomerulosclerosis, and fibrosis. Our findings underscore the potential of Dapa in preventing maladaptive repair following AKI, emphasizing the crucial role of early intervention in mitigating AKI long-term consequences.

Sulforaphane protects from kidney damage during the release of unilateral ureteral obstruction (RUUO) by activating nuclear factor erythroid 2-related factor 2 (Nrf2): Role of antioxidant, anti-inflammatory, and antiapoptotic mechanisms.

Releasing unilateral ureteral obstruction (RUUO) is the gold standard for decreasing renal damage induced during unilateral ureteral obstruction (UUO); however, the complete recovery after RUUO depends on factors such as the time and severity of obstruction and kidney contralateral compensatory mechanisms. Interestingly, previous studies have shown that kidney damage markers such as oxidative stress, inflammation, and apoptosis are present and even increase after removal obstruction. To date, previous therapeutic strategies have been used to potentiate the recovery of renal function after RUUO; however, the mechanisms involving renal damage reduction are poorly described and sometimes focus on the recovery of renal functionality. Furthermore, using natural antioxidants has not been completely studied in the RUUO model. In this study, we selected sulforaphane (SFN) because it activates the nuclear factor erythroid 2-related factor 2 (Nrf2), a transcription factor that induces an antioxidant response, decreasing oxidative stress and inflammation, preventing apoptosis. Thus, we pre-administrated SFN on the second day after UUO until day five, where we released the obstruction on the three days after UUO. Then, we assessed oxidative stress, inflammation, and apoptosis markers. Interestingly, we found that SFN administration in the RUUO model activated Nrf2, inducing its translocation to the nucleus to activate its target proteins. Thus, the Nrf2 activation upregulated glutathione (GSH) content and the antioxidant enzymes catalase, glutathione peroxidase (GPx), and glutathione reductase (GR), which reduced the oxidative stress markers. Moreover, the improvement of antioxidant response by SFN restored S-glutathionylation in the mitochondrial fraction. Activated Nrf2 also reduced inflammation by lessening the nucleotide-binding domain-like receptor family pyrin domain containing 3 and interleukin 1ß (IL-1ß) production. Reducing oxidative stress and inflammation prevented apoptosis by avoiding caspase 3 cleavage and increasing B-cell lymphoma 2 (Bcl2) levels. Taken together, the obtained results in our study showed that the upregulation of Nrf2 by SFN decreases oxidative stress, preventing inflammation and apoptosis cell death during the release of UUO.

NAC Pre-Administration Prevents Cardiac Mitochondrial Bioenergetics, Dynamics, Biogenesis, and Redox Alteration in Folic Acid-AKI-Induced Cardio-Renal Syndrome Type 3.

The incidence of kidney disease is increasing worldwide. Acute kidney injury (AKI) can strongly favor cardio-renal syndrome (CRS) type 3 development. However, the mechanism involved in CRS development is not entirely understood. In this sense, mitochondrial impairment in both organs has become a central axis in CRS physiopathology. This study aimed to elucidate the molecular mechanisms associated with cardiac mitochondrial impairment and its role in CRS development in the folic acid-induced AKI (FA-AKI) model. Our results showed that 48 h after FA-AKI, the administration of N-acetyl-cysteine (NAC), a mitochondrial glutathione regulator, prevented the early increase in inflammatory and cell death markers and oxidative stress in the heart. This was associated with the ability of NAC to protect heart mitochondrial bioenergetics, principally oxidative phosphorylation (OXPHOS) and membrane potential, through complex I activity and the preservation of glutathione balance, thus preventing mitochondrial dynamics shifting to fission and the decreases in mitochondrial biogenesis and mass. Our data show, for the first time, that mitochondrial bioenergetics impairment plays a critical role in the mechanism that leads to heart damage. Furthermore, NAC heart mitochondrial preservation during an AKI event can be a valuable strategy to prevent CRS type 3 development.

Therapeutic Effect of Curcumin on 5/6Nx Hypertriglyceridemia: Association with the Improvement of Renal Mitochondrial ß-Oxidation and Lipid Metabolism in Kidney and Liver.

Chronic kidney disease (CKD) prevalence is constantly increasing, and dyslipidemia in this disease is characteristic, favoring cardiovascular events. However, the mechanisms of CKD dyslipidemia are not fully understood. The use of curcumin (CUR) in CKD models such as 5/6 nephrectomy (5/6Nx) has shown multiple beneficial effects, so it has been proposed to correct dyslipidemia without side effects. This work aimed to characterize CUR's potential therapeutic effect on dyslipidemia and alterations in lipid metabolism and mitochondrial ß-oxidation in the liver and kidney in 5/6Nx. Male Wistar rats were subjected to 5/6Nx and progressed by 4 weeks; meanwhile, CUR (120 mg/kg) was administered for weeks 5 to 8. Our results showed that CUR reversed the increase in liver and kidney damage and hypertriglyceridemia induced by 5/6Nx. CUR also reversed mitochondrial membrane depolarization and ß-oxidation disorders in the kidney and the increased lipid uptake and the high levels of proteins involved in fatty acid synthesis in the liver and kidney. CUR also decreased lipogenesis and increased mitochondrial biogenesis markers in the liver. Therefore, we concluded that the therapeutic effect of curcumin on 5/6Nx hypertriglyceridemia is associated with the restoration of renal mitochondrial ß-oxidation and the reduction in lipid synthesis and uptake in the kidneys and liver.

Ellagic acid ameliorates hexavalent chromium-induced renal toxicity by attenuating oxidative stress, suppressing TNF-α and protecting mitochondria.

Nephrotoxicity is an important adverse effect of oxidative stress induced by hexavalent chromium [Cr(VI)]. The effect of ellagic acid, a dietary polyphenolic compound with potent antioxidant activity, was investigated in Cr(VI)-induced kidney injury. Six groups of male Wistar rats were treated intragastrically with vehicle or ellagic acid (15 and 30 mg/kg) for 10 days. On day 10, rats received saline or Cr(VI) (K2Cr2O7 15 mg/kg) subcutaneously. Cr(VI) significantly increased kidney weight, affected kidney function assessed by biomarkers in blood and urine (protein, creatinine and urea nitrogen), caused histological changes (tubular injury and glomerular capillary tuft damage), increased markers of oxidative stress and reduced the activity of antioxidant enzymes. In addition, Cr(VI) altered mitochondrial ultrastructure, impaired mitochondrial respiration, increased lipid peroxidation, and inhibited the function of mitochondrial enzymes. Pretreatment with ellagic acid (30 mg/kg) attenuated all the aforementioned alterations. Furthermore, we explored whether ellagic acid might regulate the tumor necrosis factor-alpha (TNF-α)/receptor-interacting protein kinase 3 (RIPK3) pathway, reducing Cr(VI)-induced tubular necrosis. Cr(VI) upregulated both TNF-α and RIPK3, but ellagic acid only decreased TNF-α levels, having no effect on RIPK3 content. Therefore, understanding the mechanisms through which Cr(VI) promotes necroptosis is crucial for future studies, in order to design strategies to mitigate kidney damage. In conclusion, ellagic acid attenuated Cr(VI)-induced renal alterations by preventing oxidative stress, supporting enzymatic activities, suppressing TNF-α, and preserving mitochondrial ultrastructure and function, most likely due to its antioxidant properties.

Effect of mycobacterial proteins that target mitochondria on the alveolar macrophages activation during Mycobacterium tuberculosis infection.

Purpose of the study: During the early and progressive (late) stages of murine experimental pulmonary tuberculosis, the differential activation of macrophages contributes to disease development by controlling bacterial growth and immune regulation. Mycobacterial proteins P27 and PE_PGRS33 can target the mitochondria of macrophages. This study aims to evaluate the effect of both proteins on macrophage activation during mycobacterial infection. Materials and methods: We assess both proteins for mitochondrial oxygen consumption, and morphological changes, as well as bactericide activity, production of metabolites, cytokines, and activation markers in infected MQs. The cell line MH-S was used for all the experiments. Results: We show that P27 and PE_PGRS33 proteins modified mitochondrial dynamics, oxygen consumption, bacilli growth, cytokine production, and some genes that contribute to macrophage alternative activation and mycobacterial intracellular survival. Conclusions: Our findings showed that these bacterial proteins partially contribute to promoting M2 differentiation by altering mitochondrial metabolic activity.

Quinolinic Acid Induces Alterations in Neuronal Subcellular Compartments, Blocks Autophagy Flux and Activates Necroptosis and Apoptosis in Rat Striatum.

Quinolinic acid (QUIN) is an agonist of N-methyl-D-aspartate receptor (NMDAr) used to study the underlying mechanism of excitotoxicity in animal models. There is evidence indicating that impairment in autophagy at early times contributes to cellular damage in excitotoxicity; however, the status of autophagy in QUIN model on day 7 remains unexplored. In this study, the ultrastructural analysis of subcellular compartments and the status of autophagy, necroptosis, and apoptosis in the striatum of rats administered with QUIN (120 nmol and 240 nmol) was performed on day 7. QUIN induced circling behavior, neurodegeneration, and cellular damage; also, it promoted swollen mitochondrial crests, spherical-like morphology, and mitochondrial fragmentation; decreased ribosomal density in the rough endoplasmic reticulum; and altered the continuity of myelin sheaths in axons with separation of the compact lamellae. Furthermore, QUIN induced an increase and a decrease in ULK1 and p-70-S6K phosphorylation, respectively, suggesting autophagy activation; however, the increased microtubule-associated protein 1A/1B-light chain 3-II (LC3-II) and sequestosome-1/p62 (SQSTM1/p62), the coexistence of p62 and LC3 in the same structures, and the decrease in Beclin 1 and mature cathepsin D also indicates a blockage in autophagy flux. Additionally, QUIN administration increased tumor necrosis factor alpha (TNFα) and receptor-interacting protein kinase 3 (RIPK3) levels and its phosphorylation (p-RIPK3), as well as decreased B-cell lymphoma 2 (Bcl-2) and increased Bcl-2-associated X protein (Bax) levels and c-Jun N-terminal kinase (JNK) phosphorylation, suggesting an activation of necroptosis and apoptosis, respectively. These results suggest that QUIN activates the autophagy, but on day 7, it is blocked and organelle and cellular damage, neurodegeneration, and behavior alterations could be caused by necroptosis and apoptosis activation.

Mycobacterium tuberculosis Infection Induces BCSFB Disruption but No BBB Disruption In Vivo: Implications in the Pathophysiology of Tuberculous Meningitis.

Central nervous system (CNS) tuberculosis is the most lethal and devastating form among the diseases caused by Mycobacterium tuberculosis. The mechanisms by which M. tuberculosis bacilli enter the CNS are still unclear. However, the BBB and the BCSFB have been proposed as possible routes of access into the brain. We previously reported that certain strains of M. tuberculosis possess an enhanced ability to cause secondary CNS infection in a mouse model of progressive pulmonary tuberculosis. Here, we evaluated the morphostructural and molecular integrity of CNS barriers. For this purpose, we analyzed through transmission electron microscopy the ultrastructure of brain parenchymal microvessels and choroid plexus epithelium from animals infected with two mycobacterial strains. Additionally, we determined the expression of junctional proteins and cytokines by immunological techniques. The results showed that the presence of M. tuberculosis induced disruption of the BCSFB but no disruption of the BBB, and that the severity of such damage was related to the strain used, suggesting that variations in the ability to cause CNS disease among distinct strains of bacteria may also be linked to their capacity to cause direct or indirect disruption of these barriers. Understanding the pathophysiological mechanisms involved in CNS tuberculosis may facilitate the establishment of new biomarkers and therapeutic targets.

Klf10 favors Mycobacterium tuberculosis survival by impairing IFN-γ production and preventing macrophages reprograming to macropinocytosis.

Mycobacterium tuberculosis has developed diverse mechanisms to survive inside phagocytic cells, such as macrophages. Phagocytosis is a key process in eliminating invading pathogens; thus, M. tuberculosis efficiently disrupts phagosome maturation to ensure infection. However, inflammatory cytokines produced by macrophages in response to early M. tuberculosis infection are key to promoting bacterial clarification. IFN-γ enhances M. tuberculosis engulfment and destruction by reprogramming macrophages from phagocytosis to macropinocytosis. Here, we show that the transcription factor Krüppel-like factor 10 (Klf10) plays a positive role in M. tuberculosis survival and infection by negatively modulating IFN-γ levels. Naïve Klf10-deficient macrophages produce more IFN-γ upon stimulation than wild-type macrophages, thus enhancing bacterial uptake and bactericidal activity achieved by macropinocytosis. Moreover, Klf10⁻/ ⁻ macrophages showed cytoplasmic distribution of coronin 1 correlated with increased pseudopod count and length. In agreement with these observations, Klf10⁻/ ⁻ mice showed improved bacterial clearance from the lungs and increased viability. Altogether, our data indicate that Klf10 plays a critical role in M. tuberculosis survival by preventing macrophage reprogramming from phagocytosis to macropinocytosis by negatively regulating IFN-γ production upon macrophage infection.

A ketogenic diet attenuates acute and chronic ischemic kidney injury and reduces markers of oxidative stress and inflammation.

BACKGROUND: Ischemic kidney injury is a common clinical condition resulting from transient interruption of the kidney's normal blood flow, leading to oxidative stress, inflammation, and kidney dysfunction. The ketogenic diet (KD), a low-carbohydrate, high-fat diet that stimulates endogenous ketone body production, has potent antioxidant and anti-inflammatory effects in distinct tissues and might thus protect the kidney against ischemia and reperfusion (IR) injury. MAIN METHODS: Male Wistar rats were fed a KD or a control diet (CD) for three days before analyzing metabolic parameters or testing nephroprotection. We used two different models of kidney IR injury and conducted biochemical, histological, and Western blot analyses at 24 h and two weeks after surgery. KEY FINDINGS: Acute KD feeding caused protein acetylation, liver AMPK activation, and increased resistance to IR-induced kidney injury. At 24 h after IR, rats on KD presented reduced tubular damage and improved kidney functioning compared to rats fed with a CD. KD attenuated oxidative damage (protein nitration, 4-HNE adducts, and 8-OHdG), increased antioxidant defenses (GPx and SOD activity), and reduced inflammatory intermediates (IL6, TNFα, MCP1), p50 NF-κB expression, and cellular infiltration. Also, KD prevented interstitial fibrosis development at two weeks, up-regulation of HSP70, and chronic Klotho deficiency. SIGNIFICANCE: Our findings demonstrate for the first time that short-term KD increases tolerance to experimental kidney ischemia, opening the opportunity for future therapeutic exploration of a dietary preconditioning strategy to convey kidney protection in the clinic.

In Vitro, In Vivo and In Silico Assessment of the Antimicrobial and Immunomodulatory Effects of a Water Buffalo Cathelicidin (WBCATH) in Experimental Pulmonary Tuberculosis.

Tuberculosis (TB) is considered the oldest pandemic in human history. The emergence of multidrug-resistant (MDR) strains is currently considered a serious global health problem. As components of the innate immune response, antimicrobial peptides (AMPs) such as cathelicidins have been proposed to have efficacious antimicrobial activity against Mycobacterium tuberculosis (Mtb). In this work, we assessed a cathelicidin from water buffalo, Bubalus bubalis, (WBCATH), determining in vitro its antitubercular activity (MIC), cytotoxicity and the peptide effect on bacillary loads and cytokines production in infected alveolar macrophages. Our results showed that WBCATH has microbicidal activity against drug-sensitive and MDR Mtb, induces structural mycobacterial damage demonstrated by electron microscopy, improves Mtb killing and induces the production of protective cytokines by murine macrophages. Furthermore, in vivo WBCATH showed decreased bacterial loads in a model of progressive pulmonary TB in BALB/c mice infected with drug-sensitive or MDR mycobacteria. In addition, a synergistic therapeutic effect was observed when first-line antibiotics were administered with WBCATH. These results were supported by computational modeling of the potential effects of WBCATH on the cellular membrane of Mtb. Thus, this water buffalo-derived cathelicidin could be a promising adjuvant therapy for current anti-TB drugs by enhancing a protective immune response and potentially reducing antibiotic treatment duration.

[Lymphoma with signet ring morphology: pathology with distinctive differential diagnoses]. / Linfoma con morfología en anillo de sello: patología con distintivos diagnósticos diferenciales.

Background: Signet-ring cell lymphoma is a rare entity that simulates epithelial neoplasms, sarcomas and reactive histiocytes conditions. It represents a morphological variant of non-Hodgkin's lymphomas, its diagnosis can represent a challenge, therefore it should be considered in complementary studies. Objective: The aim of this work is to show a case with a very rare morphology and to emphasize the importance of awareness this entity and avoid mistakes in its diagnosis. Clinical case: We present the case of a 67-year-old man, who developed lymph node growths in the right armpit, neck, right groin, and submandibular region, with apparent involvement of the lungs and spleen; was diagnosed as diffuse large B cell lymphoma with signet-ring morphology, originated in the germinal center. Transmission electron microscopy study was carried out for a more precise characterization of the morphology. Unfortunately, the patient did not return for a follow-up consultation, so he did not start treatment and died 6 months after diagnosis. Conclusions: Lymphoma with the signet-ring phenotype is rare, and can occur in any type of non-Hodgkin lymphoma; however, this morphology is more commonly associated with carcinomas and, less frequently, with sarcomas, melanomas or reactive histiocytes conditions, therefore should be considered this entity together with the appropriate use of complementary studies for proper diagnosis.

Introducción: el linfoma con células en anillo de sello es una entidad poco frecuente y simuladora de neoplasias epiteliales, sarcomas y condiciones reactivas de histiocitos. Representa una variante morfológica de distintos linfomas no Hodgkin, por lo que su diagnóstico puede representar un desafío y debe ser considerado al realizar estudios complementarios. Objetivo: mostrar un caso con una morfología muy poco frecuente y recalcar la importancia de conocer esta entidad para no cometer errores en su diagnóstico. Caso clínico: se documenta el caso de un hombre de 67 años que desarrolló crecimientos ganglionares en axila derecha, cuello, ingle derecha, y región submandibular, con aparente afección en pulmones y bazo, que fue diagnosticado como linfoma B difuso de células grandes con morfología en anillo de sello, originado en el centro germinal. Se realizó estudio de microscopía electrónica de transmisión para una mejor caracterización de la morfología. Desafortunadamente el paciente no regresó a consulta de seguimiento, por lo que no inició tratamiento, falleció a los 6 meses posteriores al diagnóstico. Conclusiones: el linfoma con fenotipo en anillo de sello es poco frecuente, y puede presentarse en cualquier tipo de linfomas no Hodgkin; sin embargo, esta morfología es más comúnmente asociada a carcinomas y, en menor frecuencia, a sarcomas, melanomas o histiocitos reactivos, por lo que el considerar esta entidad junto con el uso adecuado de estudios complementarios es de gran importancia para su adecuado diagnóstico.

Linfoma con morfología en anillo de sello: patología con distintivos diagnósticos diferenciales / Lymphoma with signet ring morphology: pathology with distinctive differential diagnoses

Introducción: el linfoma con células en anillo de sello es una entidad poco frecuente y simuladora de neoplasias epiteliales, sarcomas y condiciones reactivas de histiocitos. Representa una variante morfológica de distintos linfomas no Hodgkin, por lo que su diagnóstico puede representar un desafío y debe ser considerado al realizar estudios complementarios. Objetivo: mostrar un caso con una morfología muy poco frecuente y recalcar la importancia de conocer esta entidad para no cometer errores en su diagnóstico. Caso clínico: se documenta el caso de un hombre de 67 años que desarrolló crecimientos ganglionares en axila derecha, cuello, ingle derecha, y región submandibular, con aparente afección en pulmones y bazo, que fue diagnosticado como linfoma B difuso de células grandes con morfología en anillo de sello, originado en el centro germinal. Se realizó estudio de microscopía electrónica de transmisión para una mejor caracterización de la morfología. Desafortunadamente el paciente no regresó a consulta de seguimiento, por lo que no inició tratamiento, falleció a los 6 meses posteriores al diagnóstico. Conclusiones: el linfoma con fenotipo en anillo de sello es poco frecuente, y puede presentarse en cualquier tipo de linfomas no Hodgkin; sin embargo, esta morfología es más comúnmente asociada a carcinomas y, en menor frecuencia, a sarcomas, melanomas o histiocitos reactivos, por lo que el considerar esta entidad junto con el uso adecuado de estudios complementarios es de gran importancia para su adecuado diagnóstico.

Background: Signet-ring cell lymphoma is a rare entity that simulates epithelial neoplasms, sarcomas and reactive histiocytes conditions. It represents a morphological variant of non-Hodgkin's lymphomas, its diagnosis can represent a challenge, therefore it should be considered in complementary studies. Objective: The aim of this work is to show a case with a very rare morphology and to emphasize the importance of awareness this entity and avoid mistakes in its diagnosis. Clinical case: We present a case of a 67-year-old man, who developed lymph node growths in the right armpit, neck, right groin, and submandibular region, with apparent involvement of the lungs and spleen; was diagnosed as diffuse large B cell lymphoma with signet-ring morphology, originated in the germinal center. Transmission electron microscopy study was carried out for a more precise characterization of the morphology. Unfortunately, the patient did not return for a follow-up consultation, so he did not start treatment and died 6 months after diagnosis. Conclusions: Lymphoma with the signet-ring phenotype is rare, and can occur in any type of non-Hodgkin lymphoma; however, this morphology is more commonly associated with carcinomas and, less frequently, with sarcomas, melanomas or reactive histiocytes conditions, therefore should be considered this entity together with the appropriate use of complementary studies for proper diagnosis.

Progressive Reduction in Mitochondrial Mass Is Triggered by Alterations in Mitochondrial Biogenesis and Dynamics in Chronic Kidney Disease Induced by 5/6 Nephrectomy.

The five-sixth nephrectomy (5/6Nx) model is widely used to study the mechanisms involved in chronic kidney disease (CKD) progression. Mitochondrial impairment is a critical mechanism that favors CKD progression. However, until now, there are no temporal studies of the change in mitochondrial biogenesis and dynamics that allow determining the role of these processes in mitochondrial impairment and renal damage progression in the 5/6Nx model. In this work, we determined the changes in mitochondrial biogenesis and dynamics markers in remnant renal mass from days 2 to 28 after 5/6Nx. Our results show a progressive reduction in mitochondrial biogenesis triggered by reducing two principal regulators of mitochondrial protein expression, the peroxisome proliferator-activated receptor-gamma coactivator 1-alpha and the peroxisome proliferator-activated receptor alpha. Furthermore, the reduction in mitochondrial biogenesis proteins strongly correlates with the increase in renal damage markers. Additionally, we found a slow and gradual change in mitochondrial dynamics from fusion to fission, favoring mitochondrial fragmentation at later stages after 5/6Nx. Together, our results suggest that 5/6Nx induces the progressive reduction in mitochondrial mass over time via the decrease in mitochondrial biogenesis factors and a slow shift from mitochondrial fission to fusion; both mechanisms favor CKD progression in the remnant renal mass.

Quinacrine, an Antimalarial Drug with Strong Activity Inhibiting SARS-CoV-2 Viral Replication In Vitro.

Quinacrine (Qx), a molecule used as an antimalarial, has shown anticancer, antiprion, and antiviral activity. The most relevant antiviral activities of Qx are related to its ability to raise pH in acidic organelles, diminishing viral enzymatic activity for viral cell entry, and its ability to bind to viral DNA and RNA. Moreover, Qx has been used as an immunomodulator in cutaneous lupus erythematosus and various rheumatological diseases, by inhibiting phospholipase A2 modulating the Th1/Th2 response. The aim of this study was to evaluate the potential antiviral effect of Qx against denominated severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection in Vero E6 cells. The cytotoxicity of Qx in Vero E6 cells was determined by the MTT assay. Afterwards, Vero E6 cells were infected with SARS-CoV-2 at different multiplicities of infections (MOIs) of 0.1 and 0.01 in the presence of Qx (0-30 µM) to determinate the half maximal effective concentration (EC50). After 48 h, the effect of Qx against SARS-CoV-2 was assessed by viral cytotoxicity and viral copy numbers, the last were determined by digital real-time RT-PCR (ddRT-PCR). Additionally, electron and confocal microscopy of Vero E6 cells infected and treated with Qx was studied. Our data show that Qx reduces SARS-CoV-2 virus replication and virus cytotoxicity, apparently by inhibition of viral ensemble, as observed by ultrastructural images, suggesting that Qx could be a potential drug for further clinical studies against coronavirus disease 2019 (COVID-19) infection.

Antimicrobial Peptide against Mycobacterium Tuberculosis That Activates Autophagy Is an Effective Treatment for Tuberculosis.

Mycobacterium tuberculosis (MTB) is the principal cause of human tuberculosis (TB), which is a serious health problem worldwide. The development of innovative therapeutic modalities to treat TB is mainly due to the emergence of multi drug resistant (MDR) TB. Autophagy is a cell-host defense process. Previous studies have reported that autophagy-activating agents eliminate intracellular MDR MTB. Thus, combining a direct antibiotic activity against circulating bacteria with autophagy activation to eliminate bacteria residing inside cells could treat MDR TB. We show that the synthetic peptide, IP-1 (KFLNRFWHWLQLKPGQPMY), induced autophagy in HEK293T cells and macrophages at a low dose (10 µM), while increasing the dose (50 µM) induced cell death; IP-1 induced the secretion of TNFα in macrophages and killed Mtb at a dose where macrophages are not killed by IP-1. Moreover, IP-1 showed significant therapeutic activity in a mice model of progressive pulmonary TB. In terms of the mechanism of action, IP-1 sequesters ATP in vitro and inside living cells. Thus, IP-1 is the first antimicrobial peptide that eliminates MDR MTB infection by combining four activities: reducing ATP levels, bactericidal activity, autophagy activation, and TNFα secretion.

Activation of STAT3 Regulates Reactive Astrogliosis and Neuronal Death Induced by AßO Neurotoxicity.

Amyloid-beta oligomers (AßO) have been proposed as the most potent neurotoxic and inflammation inducers in Alzheimer's disease (AD). AßO contribute to AD pathogenesis by impairing the production of several cytokines and inflammation-related signaling pathways, such as the Janus kinases/signal transducer of transcription factor-3 (JAK/STAT3) pathway. STAT3 modulates glial activation, indirectly regulates Aß deposition, and induces cognitive decline in AD transgenic models. However, in vivo studies using an AßO microinjection rat model have not yet explored STAT3 role. The main purpose of this study was to elucidate if a single microinjection of AßO could promote an increased expression of STAT3 in glial cells favoring neuroinflammation and neurodegeneration. We designed a model of intrahippocampal microinjection and assessed glial activation, cytokines production, STAT3 expression, and neurodegeneration in time. Our results showed robust expression of STAT3 in glial cells (mainly in astrocytes) and neurons, correlating with neuronal death in response to AßO administration. A STAT3 inhibition assay conducted in rat primary hippocampal cultures, suggested that the induction of the transcription factor by AßO in astrocytes leads them to an activation state that may favor neuronal death. Notwithstanding, pharmacological inhibition of the JAK2/STAT3 pathway should be focused on astrocytes because it is also essential in neurons survival. Overall, these findings strongly suggest the participation of STAT3 in the development of neurodegeneration.

Temporal Alterations in Mitochondrial ß-Oxidation and Oxidative Stress Aggravate Chronic Kidney Disease Development in 5/6 Nephrectomy Induced Renal Damage.

Five-sixths nephrectomy (5/6Nx) model is widely used for studying the mechanisms involved in chronic kidney disease (CKD) progression, a kidney pathology that has increased dramatically in recent years. Mitochondrial impairment is a key mechanism that aggravates CKD progression; however, the information on mitochondrial bioenergetics and redox alterations along a time course in a 5/6Nx model is still limited and in some cases contradictory. Therefore, we performed for the first time a time-course study of mitochondrial alterations by high-resolution respirometry in the 5/6Nx model. Our results show a decrease in mitochondrial ß-oxidation at early times, as well as a permanent impairment in adenosine triphosphate (ATP) production in CI-linked respiration, a permanent oxidative state in mitochondria and decoupling of these organelles. These pathological alterations are linked to the early decrease in complex I and ATP synthase activities and to the further decrease in complex III activity. Therefore, our results may suggest that mitochondrial bioenergetics impairment is an early event in renal damage, whose persistence in time aggravates CKD development in the 5/6Nx model.

Alterations in mitochondrial homeostasis in a potassium dichromate model of acute kidney injury and their mitigation by curcumin.

Curcumin has protective effects in several acute kidney injury models, including that induced by potassium dichromate (K2Cr2O7). The protective effect of curcumin in this experimental model has been associated to the preservation of mitochondrial bioenergetics. This study is aimed at evaluating whether or not curcumin's protective effect in mitochondrial bioenergetics is related to the modulation of mitochondrial dynamics and biogenesis. Wistar rats were treated with a single subcutaneous dose of K2Cr2O7 (12.5 mg/kg) or received curcumin (400 mg/kg/day) by oral gavage 10 days before and one day after the K2Cr2O7 injection. K2Cr2O7 induced kidney dysfunction and increased mitochondrial hydrogen peroxide production, while decreasing the respiration directly attributable to oxidative phosphorylation and mitochondrial membrane potential. In mitochondria, K2Cr2O7 increased fission and reduced fusion. Structural analysis of mitochondria in the proximal tubular cells corroborated their fragmentation and loss of crests' integrity. Regarding mitochondrial biogenesis, K2Cr2O7 decreased peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α) levels. Conversely, curcumin treatment mitigated the aforementioned alterations and increased the expression of the mitochondrial transcription factor A (TFAM). Taken together, our results suggest that curcumin can protect against renal injury by modulating mitochondrial homeostasis, mitigating alterations in bioenergetics and dynamics, possibly by stimulating mitochondrial biogenesis.