International Journal of Molecular Sciences

Research

24 pages, 3397 KiB

Open AccessArticle

A Preliminary Investigation of the Roles of Endometrial Cells in Endometriosis Development via In Vitro and In Vivo Analyses

by Yin-Hua Cheng, Ching-Wei Huang, Hao-Ting Lien, Yu-Yang Hsiao, Pei-Ling Weng, Yung-Chiao Chang, Jai-Hong Cheng and Kuo-Chung Lan

Int. J. Mol. Sci. 2024, 25(7), 3873; https://doi.org/10.3390/ijms25073873 - 30 Mar 2024

Viewed by 742

Abstract

Endometriosis is a complex gynecological disease that affects more than 10% of women in their reproductive years. While surgery can provide temporary relief from women’s pain, symptoms often return in as many as 75% of cases within two years. Previous literature has contributed [...] Read more.

Endometriosis is a complex gynecological disease that affects more than 10% of women in their reproductive years. While surgery can provide temporary relief from women’s pain, symptoms often return in as many as 75% of cases within two years. Previous literature has contributed to theories about the development of endometriosis; however, the exact pathogenesis and etiology remain elusive. We conducted a preliminary investigation into the influence of primary endometrial cells (ECs) on the development and progression of endometriosis. In vitro studies, they were involved in inducing Lipopolysaccharide (LPS) in rat-isolated primary endometrial cells, which resulted in increased nuclear factor-kappa B (NF-κB) and vascular endothelial growth factor (VEGF) mRNA gene expression (quantitative polymerase chain reaction analysis, qPCR) and protein expression (western blot analysis). Additionally, in vivo studies utilized autogenic and allogeneic transplantations (rat to rat) to investigate endometriosis-like lesion cyst size, body weight, protein levels (immunohistochemistry), and mRNA gene expression. These studies demonstrated that estrogen upregulates the gene and protein regulation of cytoskeletal (CK)-18, transforming growth factor-β (TGF-β), VEGF, and tumor necrosis factor (TNF)-α, particularly in the peritoneum. These findings may influence cell proliferation, angiogenesis, fibrosis, and inflammation markers. Consequently, this could exacerbate the occurrence and progression of endometriosis. Full article

(This article belongs to the Special Issue Advances in Animal Models in Biomedical Research)

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18 pages, 29244 KiB

Open AccessArticle

The Effects of Aging on Sarcoplasmic Reticulum-Related Factors in the Skeletal Muscle of Mice

by Yuji Kanazawa, Tatsuo Takahashi, Mamoru Nagano, Satoshi Koinuma and Yasufumi Shigeyoshi

Int. J. Mol. Sci. 2024, 25(4), 2148; https://doi.org/10.3390/ijms25042148 - 10 Feb 2024

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Abstract

The pathogenesis of sarcopenia includes the dysfunction of calcium homeostasis associated with the sarcoplasmic reticulum; however, the localization in sarcoplasmic reticulum-related factors and differences by myofiber type remain unclear. Here, we investigated the effects of aging on sarcoplasmic reticulum-related factors in the soleus [...] Read more.

The pathogenesis of sarcopenia includes the dysfunction of calcium homeostasis associated with the sarcoplasmic reticulum; however, the localization in sarcoplasmic reticulum-related factors and differences by myofiber type remain unclear. Here, we investigated the effects of aging on sarcoplasmic reticulum-related factors in the soleus (slow-twitch) and gastrocnemius (fast-twitch) muscles of 3- and 24-month-old male C57BL/6J mice. There were no notable differences in the skeletal muscle weight of these 3- and 24-month-old mice. The expression of Atp2a1, Atp2a2, Sln, and Pln increased with age in the gastrocnemius muscles, but not in the soleus muscles. Subsequently, immunohistochemical analysis revealed ectopic sarcoplasmic reticulum calcium ion ATPase (SERCA) 1 and SERCA2a immunoreactivity only in the gastrocnemius muscles of old mice. Histochemical and transmission electron microscope analysis identified tubular aggregate (TA), an aggregation of the sarcoplasmic reticulum, in the gastrocnemius muscles of old mice. Dihydropyridine receptor α1, ryanodine receptor 1, junctophilin (JPH) 1, and JPH2, which contribute to sarcoplasmic reticulum function, were also localized in or around the TA. Furthermore, JPH1 and JPH2 co-localized with matrix metalloproteinase (MMP) 2 around the TA. These results suggest that sarcoplasmic reticulum-related factors are localized in or around TAs that occur in fast-twitch muscle with aging, but some of them might be degraded by MMP2. Full article

(This article belongs to the Special Issue Advances in Animal Models in Biomedical Research)

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23 pages, 9203 KiB

Open AccessArticle

A Novel Tendon Injury Model, Induced by Collagenase Administration Combined with a Thermo-Responsive Hydrogel in Rats, Reproduces the Pathogenesis of Human Degenerative Tendinopathy

by Laura Vidal, Maria Lopez-Garzon, Vanesa Venegas, Ingrid Vila, David Domínguez, Gil Rodas and Mario Marotta

Int. J. Mol. Sci. 2024, 25(3), 1868; https://doi.org/10.3390/ijms25031868 - 3 Feb 2024

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Abstract

Patellar tendinopathy is a common clinical problem, but its underlying pathophysiology remains poorly understood, primarily due to the absence of a representative experimental model. The most widely used method to generate such a model is collagenase injection, although this method possesses limitations. We [...] Read more.

Patellar tendinopathy is a common clinical problem, but its underlying pathophysiology remains poorly understood, primarily due to the absence of a representative experimental model. The most widely used method to generate such a model is collagenase injection, although this method possesses limitations. We developed an optimized rat model of patellar tendinopathy via the ultrasound-guided injection of collagenase mixed with a thermo-responsive Pluronic hydrogel into the patellar tendon of sixty male Wistar rats. All analyses were carried out at 3, 7, 14, 30, and 60 days post-injury. We confirmed that our rat model reproduced the pathophysiology observed in human patients through analyses of ultrasonography, histology, immunofluorescence, and biomechanical parameters. Tendons that were injured by the injection of the collagenase–Pluronic mixture exhibited a significant increase in the cross-sectional area (p < 0.01), a high degree of tissue disorganization and hypercellularity, significantly strong neovascularization (p < 0.01), important changes in the levels of types I and III collagen expression, and the organization and presence of intra-tendinous calcifications. Decreases in the maximum rupture force and stiffness were also observed. These results demonstrate that our model replicates the key features observed in human patellar tendinopathy. Collagenase is evenly distributed, as the Pluronic hydrogel prevents its leakage and thus, damage to surrounding tissues. Therefore, this model is valuable for testing new treatments for patellar tendinopathy. Full article

(This article belongs to the Special Issue Advances in Animal Models in Biomedical Research)

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19 pages, 5910 KiB

Open AccessArticle

A Novel Rabbit Model of Retained Hemothorax with Pleural Organization

by Christian J. De Vera, Rebekah L. Emerine, René A. Girard, Krishna Sarva, Jincy Jacob, Ali O. Azghani, Jon M. Florence, Alan Cook, Scott Norwood, Karan P. Singh, Andrey A. Komissarov, Galina Florova and Steven Idell

Int. J. Mol. Sci. 2024, 25(1), 470; https://doi.org/10.3390/ijms25010470 - 29 Dec 2023

Viewed by 786

Abstract

Retained hemothorax (RH) is a commonly encountered and potentially severe complication of intrapleural bleeding that can organize with lung restriction. Early surgical intervention and intrapleural fibrinolytic therapy have been advocated. However, the lack of a reliable, cost-effective model amenable to interventional testing has [...] Read more.

Retained hemothorax (RH) is a commonly encountered and potentially severe complication of intrapleural bleeding that can organize with lung restriction. Early surgical intervention and intrapleural fibrinolytic therapy have been advocated. However, the lack of a reliable, cost-effective model amenable to interventional testing has hampered our understanding of the role of pharmacological interventions in RH management. Here, we report the development of a new RH model in rabbits. RH was induced by sequential administration of up to three doses of recalcified citrated homologous rabbit donor blood plus thrombin via a chest tube. RH at 4, 7, and 10 days post-induction (RH4, RH7, and RH10, respectively) was characterized by clot retention, intrapleural organization, and increased pleural rind, similar to that of clinical RH. Clinical imaging techniques such as ultrasonography and computed tomography (CT) revealed the dynamic formation and resorption of intrapleural clots over time and the resulting lung restriction. RH7 and RH10 were evaluated in young (3 mo) animals of both sexes. The RH7 recapitulated the most clinically relevant RH attributes; therefore, we used this model further to evaluate the effect of age on RH development. Sanguineous pleural fluids (PFs) in the model were generally small and variably detected among different models. The rabbit model PFs exhibited a proinflammatory response reminiscent of human hemothorax PFs. Overall, RH7 results in the consistent formation of durable intrapleural clots, pleural adhesions, pleural thickening, and lung restriction. Protracted chest tube placement over 7 d was achieved, enabling direct intrapleural access for sampling and treatment. The model, particularly RH7, is amenable to testing new intrapleural pharmacologic interventions, including iterations of currently used empirically dosed agents or new candidates designed to safely and more effectively clear RH. Full article

(This article belongs to the Special Issue Advances in Animal Models in Biomedical Research)

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11 pages, 3417 KiB

Open AccessArticle

The Effect of Mice Adaptation Process on the Pathogenicity of Influenza A/South Africa/3626/2013 (H1N1)pdm09 Model Strain

by Mohammad Al Farroukh, Irina Kiseleva, Ekaterina Stepanova, Ekaterina Bazhenova, Elena Krutikova, Artem Tkachev, Anna Chistyakova, Andrey Rekstin, Ludmila Puchkova and Larisa Rudenko

Int. J. Mol. Sci. 2023, 24(24), 17386; https://doi.org/10.3390/ijms242417386 - 12 Dec 2023

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Abstract

Influenza virus strain A/South Africa/3626/2013 (H1N1)pdm09 (SA-WT) is a non-mouse-adapted model strain that has naturally high pathogenic properties in mice. It has been suggested that the high pathogenicity of this strain for mice could be due to the three strain-specific substitutions in the [...] Read more.

Influenza virus strain A/South Africa/3626/2013 (H1N1)pdm09 (SA-WT) is a non-mouse-adapted model strain that has naturally high pathogenic properties in mice. It has been suggested that the high pathogenicity of this strain for mice could be due to the three strain-specific substitutions in the polymerase complex (Q687R in PB1, N102T in PB2, and E358E/K heterogeneity in PB2). To evaluate the role of these replacements, SA-WT was passaged five times in mouse lungs, and the genome of the mouse-adapted version of the SA-WT strain (SA-M5) was sequenced. SA-M5 lost E358E/K heterogeneity and retained E358, which is the prevalent amino acid at this position among H1N1pdm09 strains. In addition, in the hemagglutinin of SA-M5, two heterogeneous substitutions (G155G/E and S190S/R) were identified. Both viruses, SA-M5 and SA-WT, were compared for their toxicity, ability to replicate, pathogenicity, and immunogenicity in mice. In mice infected with SA-M5 or SA-WT strains, toxicity, virus titer in pulmonary homogenates, and mouse survival did not differ significantly. In contrast, an increase in the immunogenicity of SA-M5 compared to SA-WT was observed. This increase could be due to the substitutions G155G/E and S190S/R in the HA of SA-M5. The prospects for using SA-M5 in studying the immunogenicity mechanisms were also discussed. Full article

(This article belongs to the Special Issue Advances in Animal Models in Biomedical Research)

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19 pages, 11236 KiB

Open AccessArticle

Tff3 Deficiency Differentially Affects the Morphology of Male and Female Intestines in a Long-Term High-Fat-Diet-Fed Mouse Model

by Kate Šešelja, Iva Bazina, Milka Vrecl, Jessica Farger, Martin Schicht, Friedrich Paulsen, Mirela Baus Lončar and Tatjana Pirman

Int. J. Mol. Sci. 2023, 24(22), 16342; https://doi.org/10.3390/ijms242216342 - 15 Nov 2023

Viewed by 843

Abstract

Trefoil factor family protein 3 (Tff3) protects the gastrointestinal mucosa and has a complex mode of action in different tissues. Here, we aimed to determine the effect of Tff3 deficiency on intestinal tissues in a long-term high-fat-diet (HFD)-fed model. A novel congenic strain [...] Read more.

Trefoil factor family protein 3 (Tff3) protects the gastrointestinal mucosa and has a complex mode of action in different tissues. Here, we aimed to determine the effect of Tff3 deficiency on intestinal tissues in a long-term high-fat-diet (HFD)-fed model. A novel congenic strain without additional metabolically relevant mutations (Tff3-/-/C57Bl6NCrl strain, male and female) was used. Wild type (Wt) and Tff3-deficient mice of both sexes were fed a HFD for 36 weeks. Long-term feeding of a HFD induces different effects on the intestinal structure of Tff3-deficient male and female mice. For the first time, we found sex-specific differences in duodenal morphology. HFD feeding reduced microvilli height in Tff3-deficient females compared to that in Wt females, suggesting a possible effect on microvillar actin filament dynamics. These changes could not be attributed to genes involved in ER and oxidative stress, apoptosis, or inflammation. Tff3-deficient males exhibited a reduced cecal crypt depth compared to that of Wt males, but this was not the case in females. Microbiome-related short-chain fatty acid content was not affected by Tff3 deficiency in HFD-fed male or female mice. Sex-related differences due to Tff3 deficiency imply the need to consider both sexes in future studies on the role of Tff in intestinal function. Full article

(This article belongs to the Special Issue Advances in Animal Models in Biomedical Research)

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19 pages, 3327 KiB

Open AccessArticle

Critical Velocity, Maximal Lactate Steady State, and Muscle MCT1 and MCT4 after Exhaustive Running in Mice

by Juan B. Orsi, Lara S. Araujo, Pedro P. M. Scariot, Emanuel E. C. Polisel, Luisa O. Cardoso, Claudio A. Gobatto and Fúlvia B. Manchado-Gobatto

Int. J. Mol. Sci. 2023, 24(21), 15753; https://doi.org/10.3390/ijms242115753 - 30 Oct 2023

Viewed by 951

Abstract

Although the critical velocity (CV) protocol has been used to determine the aerobic capacity in rodents, there is a lack of studies that compare CV with maximal lactate steady state intensity (iMLSS) in mice. As a consequence, their physiological and molecular responses after [...] Read more.

Although the critical velocity (CV) protocol has been used to determine the aerobic capacity in rodents, there is a lack of studies that compare CV with maximal lactate steady state intensity (iMLSS) in mice. As a consequence, their physiological and molecular responses after exercise until exhaustion at CV intensity remain unclear. Thus, we aimed to compare and correlate CV with iMLSS in running mice, following different mathematical models for CV estimation. We also evaluated their physiological responses and muscle MCT1 and MCT4 after running until exhaustion at CV. Thirty C57BL/6J mice were divided into two groups (exercised-E and control-C). Group E was submitted to a CV protocol (4 days), using linear (lin1 and lin2) and hyperbolic (hyp) mathematical models to determine the distance, velocity, and time to exhaustion (tlim) of each predictive CV trial, followed by an MLSS protocol. After a running effort until exhaustion at CV intensity, the mice were immediately euthanized, while group C was euthanized at rest. No differences were observed between iMLSS (21.1 ± 1.1 m.min⁻¹) and CV estimated by lin1 (21.0 ± 0.9 m.min⁻¹, p = 0.415), lin2 (21.3 ± 0.9 m.min⁻¹, p = 0.209), and hyp (20.6 ± 0.9 m.min⁻¹, p = 0.914). According to the results, CV was significantly correlated with iMLSS. After running until exhaustion at CV (tlim = 28.4 ± 8,29 min), group E showed lower concentrations of hepatic and gluteal glycogen than group C, but no difference in the content of MCT1 (p = 0.933) and MCT4 (p = 0.123) in soleus muscle. Significant correlations were not found between MCT1 and MCT4 and tlim at CV intensity. Our results reinforce that CV is a valid and non-invasive protocol to estimate the maximal aerobic capacity in mice and that the content of MCT1 and MCT4 was not decisive in determining the tlim at CV, at least when measured immediately after the running effort. Full article

(This article belongs to the Special Issue Advances in Animal Models in Biomedical Research)

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13 pages, 2545 KiB

Open AccessArticle

Novel Mouse Cell Lines and In Vivo Models for Human High-Grade Neuroendocrine Lung Carcinoma, Small Cell Lung Carcinoma (SCLC), and Large Cell Neuroendocrine Carcinoma (LCNEC)

by Enrique Recuero, Sara Lázaro, Corina Lorz, Ana Belén Enguita, Ramón Garcia-Escudero and Mirentxu Santos

Int. J. Mol. Sci. 2023, 24(20), 15284; https://doi.org/10.3390/ijms242015284 - 18 Oct 2023

Cited by 1 | Viewed by 1327

Abstract

There is a clear need to expand the toolkit of adequate mouse models and cell lines available for preclinical studies of high-grade neuroendocrine lung carcinoma (small cell lung carcinoma (SCLC) and large cell neuroendocrine carcinoma (LCNEC)). SCLC and LCNEC are two highly aggressive [...] Read more.

There is a clear need to expand the toolkit of adequate mouse models and cell lines available for preclinical studies of high-grade neuroendocrine lung carcinoma (small cell lung carcinoma (SCLC) and large cell neuroendocrine carcinoma (LCNEC)). SCLC and LCNEC are two highly aggressive tumor types with dismal prognoses and few therapeutic options. Currently, there is an extreme paucity of material, particularly in the case of LCNEC. Given the lack of murine cell lines and transplant models of LCNEC, the need is imperative. In this study, we generated and examined new models of LCNEC and SCLC transplantable cell lines derived from our previously developed primary mouse LCNEC and SCLC tumors. RNA-seq analysis demonstrated that our cell lines and syngeneic tumors maintained the transcriptome program from the original transgenic primary tumor and displayed strong similarities to human SCLC or LCNEC. Importantly, the SCLC transplanted cell lines showed the ability to metastasize and mimic this characteristic of the human condition. In summary, we generated mouse cell line tools that allow further basic and translational research as well as preclinical testing of new treatment strategies for SCLC and LCNEC. These tools retain important features of their human counterparts and address the lack of LCNEC disease models. Full article

(This article belongs to the Special Issue Advances in Animal Models in Biomedical Research)

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9 pages, 1196 KiB

Open AccessCommunication

A Lipid Nanoparticle-Based Method for the Generation of Liver-Specific Knockout Mice

by Sumiyo Morita, Takuro Horii, Mika Kimura, Ryosuke Kobayashi, Hiroki Tanaka, Hidetaka Akita and Izuho Hatada

Int. J. Mol. Sci. 2023, 24(18), 14299; https://doi.org/10.3390/ijms241814299 - 19 Sep 2023

Viewed by 1327

Abstract

Knockout mice are useful tools that can provide information about the normal function of genes, including their biochemical, developmental, and physiological roles. One problem associated with the generation of knockout mice is that the loss of some genes of interest produces a lethal [...] Read more.

Knockout mice are useful tools that can provide information about the normal function of genes, including their biochemical, developmental, and physiological roles. One problem associated with the generation of knockout mice is that the loss of some genes of interest produces a lethal phenotype. Therefore, the use of conditioned knockout mice, in which genes are disrupted in specific organs, is essential for the elucidation of disease pathogenesis and the verification of drug targets. In general, conditional knockout mice are produced using the Cre/loxP system; however, the production of the large numbers of Cre/flox knockout and control mice required for analysis requires substantial time and effort. Here, we describe the generation of liver-specific conditional knockout mice via the introduction of lipid nanoparticles encapsulating Cre mRNA into the liver of floxed mice. This technique does not require the production of offspring by mating floxed mice and is therefore more convenient than the conventional method. The results presented here demonstrate that the LNP-based method enables liver-specific gene knockout in a short period of time. Full article

(This article belongs to the Special Issue Advances in Animal Models in Biomedical Research)

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14 pages, 4371 KiB

Open AccessArticle

Class III Alcohol Dehydrogenase Plays a Key Role in the Onset of Alcohol-Related/-Associated Liver Disease as an S-Nitrosoglutathione Reductase in Mice

by Takeshi Haseba, Motoyo Maruyama, Toshio Akimoto, Isao Yamamoto, Midori Katsuyama and Takahisa Okuda

Int. J. Mol. Sci. 2023, 24(15), 12102; https://doi.org/10.3390/ijms241512102 - 28 Jul 2023

Viewed by 1025

Abstract

Lipid accumulation in the liver due to chronic alcohol consumption (CAC) is crucial in the development of alcohol liver disease (ALD). It is promoted by the NADH/NAD ratio increase via alcohol dehydrogenase (ADH)-dependent alcohol metabolism and lipogenesis increase via peroxisome proliferator-activated receptor γ [...] Read more.

Lipid accumulation in the liver due to chronic alcohol consumption (CAC) is crucial in the development of alcohol liver disease (ALD). It is promoted by the NADH/NAD ratio increase via alcohol dehydrogenase (ADH)-dependent alcohol metabolism and lipogenesis increase via peroxisome proliferator-activated receptor γ (PPARγ) in the liver. The transcriptional activity of PPARγ on lipogenic genes is inhibited by S-nitrosylation but activated by denitrosylation via S-nitrosoglutathione reductase (GSNOR), an enzyme identical to ADH3. Besides ADH1, ADH3 also participates in alcohol metabolism. Therefore, we investigated the specific contribution of ADH3 to ALD onset. ADH3-knockout (Adh3-/-) and wild-type (WT) mice were administered a 10% ethanol solution for 12 months. Adh3-/- exhibited no significant pathological changes in the liver, whereas WT exhibited marked hepatic lipid accumulation (p < 0.005) with increased serum transaminase levels. Adh3-/- exhibited no death during CAC, whereas WT exhibited a 40% death. Liver ADH3 mRNA levels were elevated by CAC in WT (p < 0.01). The alcohol elimination rate measured after injecting 4 g/kg ethanol was not significantly different between two strains, although the rate was increased in both strains by CAC. Thus, ADH3 plays a key role in the ALD onset, likely by acting as GSNOR. Full article

(This article belongs to the Special Issue Advances in Animal Models in Biomedical Research)

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12 pages, 2288 KiB

Open AccessArticle

Quantitative Threshold Determination of Auditory Brainstem Responses in Mouse Models

by Kenji Tanaka, Shuma Ohara, Tadaaki Matsuzaka, Aira Matsugaki, Takuya Ishimoto, Ryosuke Ozasa, Yukiko Kuroda, Koichi Matsuo and Takayoshi Nakano

Int. J. Mol. Sci. 2023, 24(14), 11393; https://doi.org/10.3390/ijms241411393 - 13 Jul 2023

Viewed by 1225

Abstract

The auditory brainstem response (ABR) is a scalp recording of potentials produced by sound stimulation, and is commonly used as an indicator of auditory function. However, the ABR threshold, which is the lowest audible sound pressure, cannot be objectively determined since it is [...] Read more.

The auditory brainstem response (ABR) is a scalp recording of potentials produced by sound stimulation, and is commonly used as an indicator of auditory function. However, the ABR threshold, which is the lowest audible sound pressure, cannot be objectively determined since it is determined visually using a measurer, and this has been a problem for several decades. Although various algorithms have been developed to objectively determine ABR thresholds, they remain lacking in terms of accuracy, efficiency, and convenience. Accordingly, we proposed an improved algorithm based on the mutual covariance at adjacent sound pressure levels. An ideal ABR waveform with clearly defined waves I–V was created; moreover, using this waveform as a standard template, the experimentally obtained ABR waveform was inspected for disturbances based on mutual covariance. The ABR testing was repeated if the value was below the established cross-covariance reference value. Our proposed method allowed more efficient objective determination of ABR thresholds and a smaller burden on experimental animals. Full article

(This article belongs to the Special Issue Advances in Animal Models in Biomedical Research)

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17 pages, 5506 KiB

Open AccessArticle

A Rat Model of Clinically Relevant Extracorporeal Circulation Develops Early Organ Dysfunctions

by Antoine Persello, Fouzia Souab, Thomas Dupas, Virginie Aillerie, Edith Bigot, Manon Denis, Angélique Erraud, Thomas Pelé, Angélique Blangy-Letheule, Pierre Miniou, Philippe Guedat, Michel De Waard, Emmanuelle Abgueguen, Bertrand Rozec and Benjamin Lauzier

Int. J. Mol. Sci. 2023, 24(8), 7338; https://doi.org/10.3390/ijms24087338 - 16 Apr 2023

Viewed by 1846

Abstract

In clinical practice, extracorporeal circulation (ECC) is associated with coagulopathy and inflammation, eventually leading to organ injuries without preventive systemic pharmacological treatment. Relevant models are needed to reproduce the pathophysiology observed in humans and preclinical tests. Rodent models are less expensive than large [...] Read more.

In clinical practice, extracorporeal circulation (ECC) is associated with coagulopathy and inflammation, eventually leading to organ injuries without preventive systemic pharmacological treatment. Relevant models are needed to reproduce the pathophysiology observed in humans and preclinical tests. Rodent models are less expensive than large models but require adaptations and validated comparisons to clinics. This study aimed to develop a rat ECC model and to establish its clinical relevance. One hour of veno-arterial ECC or a sham procedure were achieved on mechanically ventilated rats after cannulations with a mean arterial pressure objective > 60 mmHg. Five hours post-surgery, the rats’ behavior, plasmatic/blood biomarkers, and hemodynamics were measured. Blood biomarkers and transcriptomic changes were compared in 41 patients undergoing on-pump cardiac surgery. Five hours post-ECC, the rats presented hypotension, hyperlactatemia, and behavioral alterations. The same patterns of marker measurements (Lactate dehydrogenase, Creatinine kinase, ASAT, ALAT, and Troponin T) were observed in both rats and human patients. Transcriptome analyses showed similarity in both humans and rats in the biological processes involved in the ECC response. This new ECC rat model seems to resemble both ECC clinical procedures and the associated pathophysiology, but with early organ injury corresponding to a severe phenotype. Although the mechanisms at stake in the post-ECC pathophysiology of rats or humans need to be described, this new rat model appears to be a relevant and costless preclinical model of human ECC. Full article

(This article belongs to the Special Issue Advances in Animal Models in Biomedical Research)

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14 pages, 2733 KiB

Open AccessArticle

Hepatocyte Nuclear Factor-1α Increases Fibrinogen Gene Expression in Liver and Plasma Fibrinogen Concentration in Rats with Experimental Chronic Renal Failure

by Elzbieta Sucajtys-Szulc, Alicja Debska-Slizien, Boleslaw Rutkowski, Ryszard Milczarek, Marek Szolkiewicz, Julian Swierczynski and Ryszard Tomasz Smolenski

Int. J. Mol. Sci. 2023, 24(6), 5733; https://doi.org/10.3390/ijms24065733 - 17 Mar 2023

Viewed by 1409

Abstract

Chronic kidney disease (CKD) is associated with elevated plasma fibrinogen concentration. However, the underlying molecular mechanism for elevated plasma fibrinogen concentration in CKD patients has not yet been clarified. We recently found that HNF1α was significantly upregulated in the liver of chronic renal [...] Read more.

Chronic kidney disease (CKD) is associated with elevated plasma fibrinogen concentration. However, the underlying molecular mechanism for elevated plasma fibrinogen concentration in CKD patients has not yet been clarified. We recently found that HNF1α was significantly upregulated in the liver of chronic renal failure (CRF) rats, an experimental model of CKD in patients. Given that the promoter region of the fibrinogen gene possesses potential binding sites for HNF1α, we hypothesized that the upregulation of HNF1α can increase fibrinogen gene expression and consequently plasma fibrinogen concentration in the experimental model of CKD. Here, we found the coordinated upregulation of Aα-chain fibrinogen and Hnfα gene expression in the liver and elevated plasma fibrinogen concentrations in CRF rats, compared with pair-fed and control animals. Liver Aα-chain fibrinogen and HNF1α mRNAs levels correlated positively with (a) liver and plasma fibrinogen levels and (b) liver HNF1α protein levels. The positive correlation between (a) liver Aα-chain fibrinogen mRNA level, (b) liver Aα-chain fibrinogen level, and (c) serum markers of renal function suggest that fibrinogen gene transcription is closely related to the progression of kidney disease. Knockdown of Hnfα in the HepG2 cell line by small interfering RNA (siRNA) led to a decrease in fibrinogen mRNA levels. Clofibrate, an anti-lipidemic drug that reduces plasma fibrinogen concentration in humans, decreased both HNF1α and Aα-chain fibrinogen mRNAs levels in (a) the liver of CRF rats and (b) HepG2 cells. The obtained results suggest that (a) an elevated level of liver HNF1α can play an important role in the upregulation of fibrinogen gene expression in the liver of CRF rats, leading to an elevated concentration of plasma fibrinogen, a protein related to the risk of cardiovascular disease in CKD patients, and (b) fibrates can decrease plasma fibrinogen concentration through inhibition of HNF1α gene expression. Full article

(This article belongs to the Special Issue Advances in Animal Models in Biomedical Research)

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10 pages, 2850 KiB

Open AccessArticle

Outbred Mice with Streptozotocin-Induced Diabetes Show Sex Differences in Glucose Metabolism

by Boyoung Kim, Eun-Sun Park, Jong-Sun Lee and Jun-Gyo Suh

Int. J. Mol. Sci. 2023, 24(6), 5210; https://doi.org/10.3390/ijms24065210 - 8 Mar 2023

Cited by 6 | Viewed by 1445

Abstract

Outbred mice (ICR) with different genotypes and phenotypes have been reported to be more suitable for scientific testing than inbred mice because they are more similar to humans. To investigate whether the sex and genetic background of the mice are important factors in [...] Read more.

Outbred mice (ICR) with different genotypes and phenotypes have been reported to be more suitable for scientific testing than inbred mice because they are more similar to humans. To investigate whether the sex and genetic background of the mice are important factors in the development of hyperglycemia, we used ICR mice and divided them into male, female, and ovariectomized female (FOVX) groups and treated them with streptozotocin (STZ) for five consecutive days to induce diabetes. Our results show that fasting blood glucose and hemoglobin A_1c (HbA_1c) levels were significantly higher in diabetes-induced males (M-DM) and ovariectomized diabetes-induced females (FOVX-DM) than in diabetes-induced females (F-DM) at 3 and 6 weeks after STZ treatment. Furthermore, the M-DM group showed the most severe glucose tolerance, followed by the FOVX-DM and F-DM groups, suggesting that ovariectomy affects glucose tolerance in female mice. The size of pancreatic islets in the M-DM and FOVX-DM groups was significantly different from that of the F-DM group. The M-DM and FOVX-DM groups had pancreatic beta-cell dysfunction 6 weeks after STZ treatment. Urocortin 3 and somatostatin inhibited insulin secretion in the M-DM and FOVX-DM groups. Overall, our results suggest that glucose metabolism in mice is dependent on sex and/or genetic background. Full article

(This article belongs to the Special Issue Advances in Animal Models in Biomedical Research)

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11 pages, 3359 KiB

Open AccessArticle

Interplay between Learning and Voluntary Wheel Running in Male C57BL/6NCrl Mice

by Laura Niiranen, Ville Stenbäck, Mikko Tulppo, Karl-Heinz Herzig and Kari A. Mäkelä

Int. J. Mol. Sci. 2023, 24(5), 4259; https://doi.org/10.3390/ijms24054259 - 21 Feb 2023

Cited by 1 | Viewed by 1992

Abstract

Exercise is shown to improve cognitive function in various human and animal studies. Laboratory mice are often used as a model to study the effects of physical activity and running wheels provide a voluntary and non-stressful form of exercise. The aim of the [...] Read more.

Exercise is shown to improve cognitive function in various human and animal studies. Laboratory mice are often used as a model to study the effects of physical activity and running wheels provide a voluntary and non-stressful form of exercise. The aim of the study was to analyze whether the cognitive state of a mouse is related to its wheel-running behavior. Twenty-two male C57BL/6NCrl mice (9.5 weeks old) were used in the study. The cognitive function of group-housed mice (n = 5–6/group) was first analyzed in the IntelliCage system followed by individual phenotyping with the PhenoMaster with access to a voluntary running wheel. The mice were divided into three groups according to their running wheel activity: low, average, and high runners. The learning trials in the IntelliCage showed that the high-runner mice exhibited a higher error rate at the beginning of learning trials but improved their outcome and learning performance more compared to the other groups. The high-runner mice ate more compared to the other groups in the PhenoMaster analyses. There were no differences in the corticosterone levels between the groups, indicating similar stress responses. Our results demonstrate that high-runner mice exhibit enhanced learning capabilities prior to access to voluntary running wheels. In addition, our results also show that individual mice react differently when introduced to running wheels, which should be taken into consideration when choosing animals for voluntary endurance exercise studies. Full article

(This article belongs to the Special Issue Advances in Animal Models in Biomedical Research)

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24 pages, 6430 KiB

Open AccessArticle

Cardiac-Specific Expression of Cre Recombinase Leads to Age-Related Cardiac Dysfunction Associated with Tumor-like Growth of Atrial Cardiomyocyte and Ventricular Fibrosis and Ferroptosis

by Zhongguang Li, Qinchun Duan, Ying Cui, Odell D. Jones, Danyang Shao, Jianfei Zhang, Yuru Gao, Xixi Cao, Shulin Wang, Jiali Li, Xinjuan Lei, Wei Zhang, Liyang Wang, Xin Zhou, Mengmeng Xu, Yingli Liu, Jianjie Ma and Xuehong Xu

Int. J. Mol. Sci. 2023, 24(4), 3094; https://doi.org/10.3390/ijms24043094 - 4 Feb 2023

Cited by 3 | Viewed by 2710

Abstract

Transgenic expression of Cre recombinase driven by a specific promoter is normally used to conditionally knockout a gene in a tissue- or cell-type-specific manner. In αMHC-Cre transgenic mouse model, expression of Cre recombinase is controlled by the myocardial-specific α-myosin heavy chain (αMHC) promoter, [...] Read more.

Transgenic expression of Cre recombinase driven by a specific promoter is normally used to conditionally knockout a gene in a tissue- or cell-type-specific manner. In αMHC-Cre transgenic mouse model, expression of Cre recombinase is controlled by the myocardial-specific α-myosin heavy chain (αMHC) promoter, which is commonly used to edit myocardial-specific genes. Toxic effects of Cre expression have been reported, including intro-chromosome rearrangements, micronuclei formation and other forms of DNA damage, and cardiomyopathy was observed in cardiac-specific Cre transgenic mice. However, mechanisms associated with Cardiotoxicity of Cre remain poorly understood. In our study, our data unveiled that αMHC-Cre mice developed arrhythmias and died after six months progressively, and none of them survived more than one year. Histopathological examination showed that αMHC-Cre mice had aberrant proliferation of tumor-like tissue in the atrial chamber extended from and vacuolation of ventricular myocytes. Furthermore, the αMHC-Cre mice developed severe cardiac interstitial and perivascular fibrosis, accompanied by significant increase of expression levels of MMP-2 and MMP-9 in the cardiac atrium and ventricular. Moreover, cardiac-specific expression of Cre led to disintegration of the intercalated disc, along with altered proteins expression of the disc and calcium-handling abnormality. Comprehensively, we identified that the ferroptosis signaling pathway is involved in heart failure caused by cardiac-specific expression of Cre, on which oxidative stress results in cytoplasmic vacuole accumulation of lipid peroxidation on the myocardial cell membrane. Taken together, these results revealed that cardiac-specific expression of Cre recombinase can lead to atrial mesenchymal tumor-like growth in the mice, which causes cardiac dysfunction, including cardiac fibrosis, reduction of the intercalated disc and cardiomyocytes ferroptosis at the age older than six months in mice. Our study suggests that αMHC-Cre mouse models are effective in young mice, but not in old mice. Researchers need to be particularly careful when using αMHC-Cre mouse model to interpret those phenotypic impacts of gene responses. As the Cre-associated cardiac pathology matched mostly to that of the patients, the model could also be employed for investigating age-related cardiac dysfunction. Full article

(This article belongs to the Special Issue Advances in Animal Models in Biomedical Research)

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14 pages, 2613 KiB

Open AccessArticle

Half-Life Extension and Biodistribution Modulation of Biotherapeutics via Red Blood Cell Hitch-Hiking with Novel Anti-Band 3 Single-Domain Antibodies

by Toan D. Nguyen, Brandon M. Bordeau, Yu Zhang, Anna G. Mattle and Joseph P. Balthasar

Int. J. Mol. Sci. 2023, 24(1), 475; https://doi.org/10.3390/ijms24010475 - 28 Dec 2022

Cited by 4 | Viewed by 2859

Abstract

Small therapeutic proteins are receiving increased interest as therapeutic drugs; however, their clinical success has been limited due to their rapid elimination. Here, we report a half-life extension strategy via strategy via red blood cell red blood cell (RBC) hitch-hiking. This manuscript details [...] Read more.

Small therapeutic proteins are receiving increased interest as therapeutic drugs; however, their clinical success has been limited due to their rapid elimination. Here, we report a half-life extension strategy via strategy via red blood cell red blood cell (RBC) hitch-hiking. This manuscript details the development and characterization of novel anti-RBC single-domain antibodies (sdAbs), their genetic fusion to therapeutic antibody fragments (TAF) as bispecific fusion constructs, and their influence on TAF pharmacokinetics and biodistribution. Several sdAbs specific to the band 3 antigen were generated via phage-display technology. Binding affinity to RBCs was assessed via flow cytometry. Affinity maturation via random mutagenesis was carried out to improve the binding affinity of the sdAbs. Bi-specific constructs were generated by fusing the anti-RBC sdAbs with anti-tissue necrosis factor alpha (TNF-α) TAF via the use of a glycine-serine flexible linker, and assessments for binding were performed via enzyme-linked immunosorbent assay and flow cytometry. Pharmacokinetics of anti-RBC sdAbs and fusion constructs were evaluated following intravenous bolus dosing in mice at a 1 mg/kg dose. Two RBC-binding sdAbs, RB12 and RE8, were developed. These two clones showed high binding affinity to human RBC with an estimated K_D of 17.7 nM and 23.6 nM and low binding affinity to mouse RBC with an estimated K_D of 335 nM and 528 nM for RB12 and RE8, respectively. Two derivative sdAbs, RMA1, and RMC1, with higher affinities against mouse RBC, were generated via affinity maturation (K_D of 66.9 nM and 30.3 nM, respectively). Pharmacokinetic investigations in mice demonstrated prolonged circulation half-life of an anti-RBC-TNF-α bispecific construct (75 h) compared to a non-RBC binding control (1.3 h). In summary, the developed anti-RBC sdAbs and fusion constructs have demonstrated high affinity in vitro, and sufficient half-life extension in vivo. Full article

(This article belongs to the Special Issue Advances in Animal Models in Biomedical Research)

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Review

Jump to: Research, Other

15 pages, 889 KiB

Open AccessReview

Lessons from Animal Models in Sjögren’s Syndrome

by Diana Mieliauskaitė, Vilius Kontenis and Almantas Šiaurys

Int. J. Mol. Sci. 2023, 24(16), 12995; https://doi.org/10.3390/ijms241612995 - 20 Aug 2023

Cited by 1 | Viewed by 1248

Abstract

Primary Sjögren’s syndrome (pSS) is a connective tissue disease characterized by a wide spectrum of clinical features, extending from a benign glandular disease to an aggressive systemic disorder and/or lymphoma. The pathogenesis of Sjögren’s syndrome (SS) is not completely understood, but it is [...] Read more.

Primary Sjögren’s syndrome (pSS) is a connective tissue disease characterized by a wide spectrum of clinical features, extending from a benign glandular disease to an aggressive systemic disorder and/or lymphoma. The pathogenesis of Sjögren’s syndrome (SS) is not completely understood, but it is assumed that pathogenesis of SS is multifactorial. The studies based on the animal models of SS provided significant insight in SS disease pathogenesis and management. The aim of this review is to summarize current studies on animal models with primary SS-like symptoms and discuss the impact of these studies on better understanding pathogenesis and management of Sjögren’s syndrome. Databases PubMed, Web of Science, Scopus and Cochrane library were searched for summarizing studies on animal models in SS. Available data demonstrate that animal models are highly important for our understanding of SS disease. Full article

(This article belongs to the Special Issue Advances in Animal Models in Biomedical Research)

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16 pages, 2938 KiB

Open AccessReview

Preclinical Ultrasonography in Rodent Models of Neuromuscular Disorders: The State of the Art for Diagnostic and Therapeutic Applications

by Antonietta Mele, Paola Mantuano, Brigida Boccanegra, Elena Conte, Antonella Liantonio and Annamaria De Luca

Int. J. Mol. Sci. 2023, 24(5), 4976; https://doi.org/10.3390/ijms24054976 - 4 Mar 2023

Cited by 1 | Viewed by 1392

Abstract

Ultrasonography is a safe, non-invasive imaging technique used in several fields of medicine, offering the possibility to longitudinally monitor disease progression and treatment efficacy over time. This is particularly useful when a close follow-up is required, or in patients with pacemakers (not suitable [...] Read more.

Ultrasonography is a safe, non-invasive imaging technique used in several fields of medicine, offering the possibility to longitudinally monitor disease progression and treatment efficacy over time. This is particularly useful when a close follow-up is required, or in patients with pacemakers (not suitable for magnetic resonance imaging). By virtue of these advantages, ultrasonography is commonly used to detect multiple skeletal muscle structural and functional parameters in sports medicine, as well as in neuromuscular disorders, e.g., myotonic dystrophy and Duchenne muscular dystrophy (DMD). The recent development of high-resolution ultrasound devices allowed the use of this technique in preclinical settings, particularly for echocardiographic assessments that make use of specific guidelines, currently lacking for skeletal muscle measurements. In this review, we describe the state of the art for ultrasound skeletal muscle applications in preclinical studies conducted in small rodents, aiming to provide the scientific community with necessary information to support an independent validation of these procedures for the achievement of standard protocols and reference values useful in translational research on neuromuscular disorders. Full article

(This article belongs to the Special Issue Advances in Animal Models in Biomedical Research)

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21 pages, 1046 KiB

Open AccessReview

The Contribution of the Zebrafish Model to the Understanding of Polycomb Repression in Vertebrates

by Mariette Hanot, Ludivine Raby, Pamela Völkel, Xuefen Le Bourhis and Pierre-Olivier Angrand

Int. J. Mol. Sci. 2023, 24(3), 2322; https://doi.org/10.3390/ijms24032322 - 24 Jan 2023

Viewed by 1983

Abstract

Polycomb group (PcG) proteins are highly conserved proteins assembled into two major types of complexes, PRC1 and PRC2, involved in the epigenetic silencing of a wide range of gene expression programs regulating cell fate and tissue development. The crucial role of PRC1 and [...] Read more.

Polycomb group (PcG) proteins are highly conserved proteins assembled into two major types of complexes, PRC1 and PRC2, involved in the epigenetic silencing of a wide range of gene expression programs regulating cell fate and tissue development. The crucial role of PRC1 and PRC2 in the fundamental cellular processes and their involvement in human pathologies such as cancer attracted intense attention over the last few decades. Here, we review recent advancements regarding PRC1 and PRC2 function using the zebrafish model. We point out that the unique characteristics of the zebrafish model provide an exceptional opportunity to increase our knowledge of the role of the PRC1 and PRC2 complexes in tissue development, in the maintenance of organ integrity and in pathology. Full article

(This article belongs to the Special Issue Advances in Animal Models in Biomedical Research)

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Other

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12 pages, 1200 KiB

Open AccessPerspective

Dissecting the Complexity of Skeletal-Malocclusion-Associated Phenotypes: Mouse for the Rescue

by Iqbal M. Lone, Osayd Zohud, Aysar Nashef, Christian Kirschneck, Peter Proff, Nezar Watted and Fuad A. Iraqi

Int. J. Mol. Sci. 2023, 24(3), 2570; https://doi.org/10.3390/ijms24032570 - 29 Jan 2023

Cited by 12 | Viewed by 1929

Abstract

Skeletal deformities and malocclusions being heterogeneous traits, affect populations worldwide, resulting in compromised esthetics and function and reduced quality of life. Skeletal Class III prevalence is the least common of all angle malocclusion classes, with a frequency of 7.2%, while Class II prevalence [...] Read more.

Skeletal deformities and malocclusions being heterogeneous traits, affect populations worldwide, resulting in compromised esthetics and function and reduced quality of life. Skeletal Class III prevalence is the least common of all angle malocclusion classes, with a frequency of 7.2%, while Class II prevalence is approximately 27% on average, varying in different countries and between ethnic groups. Orthodontic malocclusions and skeletal deformities have multiple etiologies, often affected and underlined by environmental, genetic and social aspects. Here, we have conducted a comprehensive search throughout the published data until the time of writing this review for already reported quantitative trait loci (QTL) and genes associated with the development of skeletal deformation-associated phenotypes in different mouse models. Our search has found 72 significant QTL associated with the size of the mandible, the character, shape, centroid size and facial shape in mouse models. We propose that using the collaborative cross (CC), a highly diverse mouse reference genetic population, may offer a novel venue for identifying genetic factors as a cause for skeletal deformations, which may help to better understand Class III malocclusion-associated phenotype development in mice, which can be subsequently translated to humans. We suggest that by performing a genome-wide association study (GWAS), an epigenetics-wide association study (EWAS), RNAseq analysis, integrating GWAS and expression quantitative trait loci (eQTL), micro and small RNA, and long noncoding RNA analysis in tissues associated with skeletal deformation and Class III malocclusion characterization/phenotypes, including mandibular basic bone, gum, and jaw, in the CC mouse population, we expect to better identify genetic factors and better understand the development of this disease. Full article

(This article belongs to the Special Issue Advances in Animal Models in Biomedical Research)

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