Hepatic Sinusoids Research Articles

The α-1 Adrenergic Receptor Antagonist Doxazosin Attenuates Liver Fibrosis by Alleviating Sinusoidal Capillarization and Liver Angiogenesis.

Liver fibrosis and cirrhosis, which are caused by chronic liver injury, represent common and intractable clinical challenges of global importance. However, effective therapeutics are lacking. Therefore, the study examines the effect of doxazosin on liver fibrosis. Carbon tetrachloride (CCl4) is injected into mice to establish a liver fibrosis model. Doxazosin (5 and 10mg/kg) is administered daily by gavage. HE staining, Masson staining, Sirius Red staining, scanning electron microscopy, western blotting, real-time PCR, and immunofluorescence analysis are performed to estimate liver fibrosis and sinusoidal capillarization in mice. Cell Counting Kit-8 assays, western blotting, immunofluorescence analysis, tube formation, and transwell migration assays are performed on human umbilical vein endothelial cells (HUVECs) and human hepatic sinusoidal endothelial cells (HHSECs) to elucidate the potential mechanism of doxazosin. Doxazosin alleviates liver fibrosis and sinusoidal capillarization in CCl4-induced mice. Angiogenesis is attenuated by doxazosin in HUVECs and HHSECs. This study demonstrates that doxazosin attenuated liver fibrosis by alleviating sinusoidal capillarization and liver angiogenesis.

Neutrophil-Based Bionic Delivery System Breaks Through the Capillary Barrier of Liver Sinusoidal Endothelial Cells and Inhibits the Activation of Hepatic Stellate Cells.

The capillarization of hepatic sinusoids resulting from the activation of hepatic stellate cells poses a significant challenge, impeding the effective delivery of therapeutic agents to the Disse space for liver fibrosis treatment. Therefore, overcoming these barriers and achieving efficient drug delivery to activated hepatic stellate cells (aHSCs) are pressing challenge. In this study, we developed a synergistic sequential drug delivery approach utilizing neutrophil membrane hybrid liposome@atorvastatin/amlisentan (NCM@AtAm) and vitamin A-neutrophil membrane hybrid liposome @albumin (VNCM@Bai) nanoparticles (NPs) to breach the capillary barrier for targeted HSC cell delivery. Initially, NCM@AtAm NPs were successfully directed to the site of hepatic fibrosis through neutrophil-mediated inflammatory targeting, resulting in the normalization of liver sinusoidal endothelial cells (LSECs) and restoration of fenestrations under the combined influence of At and Am. Elevated tissue levels of the p-Akt protein and endothelial nitric oxide synthase (eNOS) indicated the normalization of LSECs following treatment with At and Am. Subsequently, VNCM@Bai NPs traversed the restored LSEC fenestrations to access the Disse space, facilitating the delivery of Bai into aHSCs under vitamin A guidance. Lastly, both in vitro and in vivo results demonstrated the efficacy of Bai in inhibiting HSC cell activation by modulating the PPAR γ/TGF-β1 and STAT1/Smad7 signaling pathways, thereby effectively treating liver fibrosis. Overall, our designed synergistic sequential delivery system effectively overcomes the barrier imposed by LSECs, offering a promising therapeutic strategy for liver fibrosis treatment in clinical settings.

Single-cell transcriptome reveals a novel mechanism of C-Kit+-liver sinusoidal endothelial cells in NASH

AimTo understand how liver sinusoidal endothelial cells (LSECs) respond to nonalcoholic steatohepatitis (NASH).MethodsWe profiled single-LSEC from livers of control and MCD-fed mice. The functions of C-Kit+-LSECs were determined using coculture and bone marrow transplantation (BMT) methods.ResultsThree special clusters of single-LSEC were differentiated. C-Kit+-LSECs of cluster 0, Msr1+-LSECs of cluster 1 and Bmp4+Selp+-VECs of cluster 2 were revealed, and these cells with diverse ectopic expressions of genes participated in regulation of endothelial, fibrosis and lipid metabolism in NASH. The number of C-Kit+-primary LSECs isolated from MCD mice was lower than control mice. Immunofluorescence co-staining of CD31 and C-KIT showed C-Kit+-LSECs located in hepatic sinusoid were also reduced in NASH patients and MCD mice, compared to AIH patients and control mice respectively. Interestingly, lipotoxic hepatocytes/HSCs cocultured with C-Kit+-LSECs or the livers of MCD mice receipting of C-Kit+-BMCs (bone marrow cells) showed less steatosis, inflammation and fibrosis, higher expression of prolipolytic FXR and PPAR-α, lower expression of TNF-α and α-SMA. Furthermore, coculturing or BMT of C-Kit+-endothelial derived cells could increase the levels of hepatic mitochondrial LC3B, decrease the degree of mitochondrial damage and ROS production through activating Pink1-mediated mitophagy pathway in NASH.ConclusionsHence, a novel transcriptomic view of LSECs was revealed to have heterogeneity and complexity in NASH. Importantly, a cluster of C-Kit+-LSECs was confirmed to recovery Pink1-related mitophagy and NASH progression.

Morphological findings in different subtypes of hepatic amyloid

The classic findings have been well described for light-chain amyloid involving the liver. In addition to light chain, however, many additional proteins are now known to be amyloidogenic and can involve the liver. A total of 58 surgical pathology specimens with amyloid deposits were analyzed for patterns of amyloid deposition, including amyloid from light chain lambda (N = 17), light chain kappa (N = 15), transthyretin (N = 15), serum amyloid A (N = 4), apolipoprotein A1 (N = 4), fibrinogen alpha (N = 2), LECT2 (N = 1). Amyloid deposits predominately targeted the liver vasculature, including the walls of the hepatic arteries, portal veins, and sinusoids. While there was overlap, light chain amyloid predominately involved the sinusoids, while transthyretin amyloid predominately targeted the hepatic arteries, especially the larger ones in the hilum and larger portal tracts. Serum amyloid A formed nodular deposits that started in the portal vasculature but then extended into the portal tract stroma, leading to large, bulbous, portal-based amyloid deposits. Apolipoprotein A amyloid also formed large portal-based nodules. Fibrinogen was mild and subtle on H&E and predominately affected portal veins. Amyloid deposits in hilar nerves were prominent with amyloid light chain, transthyretin, and apolipoprotein A1. In conclusion, the histology of hepatic amyloid is diverse and shows several distinct clusters of findings that can aide in recognition in surgical pathology specimens.

Blood defense system - Proposal for a new concept of an immune system against blood borne pathogens comprising the liver, spleen and bone marrow.

Blood-borne pathogen (BBP) infections can rapidly progress to life-threatening sepsis and must therefore be promptly eliminated by the host's immune system. Intravascular macrophages of the liver sinusoid, splenic marginal zone and red pulp and perisinusoidal macrophage protrusions in the bone marrow (BM) directly phagocytose BBPs in the blood as an innate immune response. The liver, spleen and BM thereby work together as the blood defence system (BDS) in response to BBPs by exerting their different immunological roles. The liver removes the vast majority of these invading organisms via innate immunity, but their complete elimination is not possible without the actions of antibodies. Splenic marginal zone B cells promptly produce IgM and IgG antibodies against BBPs. The splenic marginal zone transports antigenic information from the innate to the adaptive immune systems. The white pulp of the spleen functions as adaptive immune tissue and produces specific and high-affinity antibodies with an immune memory against BBPs. The BM works to maintain immune memory by supporting the survival of memory B cells, memory T cells and long-lived plasma cells (LLPCs), all of which have dedicated niches. Furthermore, BM perisinusoidal naïve follicular B cells promptly produce IgM antibodies against BBPs in the BM sinusoid and the IgG memory B cells residing in the BM rapidly transform to plasma cells which produce high-affinity IgG antibodies upon reinfection. This review describes the complete immune defence characteristics of the BDS against BBPs through the collaboration of the liver, spleen and BM with combined different immunological roles.

Consequences of Amyloid-β Deficiency for the Liver.

The hepatic content of amyloid beta (Aβ) decreases drastically in human and rodent cirrhosis highlighting the importance of understanding the consequences of Aβ deficiency in the liver. This is especially relevant in view of recent advances in anti-Aβ therapies for Alzheimer's disease (AD). Here, it is shown that partial hepatic loss of Aβ in transgenic AD mice immunized with Aβ antibody 3D6 and its absence in amyloid precursor protein (APP) knockout mice (APP-KO), as well as in human liver spheroids with APP knockdown upregulates classical hallmarks of fibrosis, smooth muscle alpha-actin, and collagen type I. Aβ absence in APP-KO and deficiency in immunized mice lead to strong activation of transforming growth factor-β (TGFβ), alpha secretases, NOTCH pathway, inflammation, decreased permeability of liver sinusoids, and epithelial-mesenchymal transition. Inversely, increased systemic and intrahepatic levels of Aβ42 in transgenic AD mice and neprilysin inhibitor LBQ657-treated wild-type mice protect the liver against carbon tetrachloride (CCl4)-induced injury. Transcriptomic analysis of CCl4-treated transgenic AD mouse livers uncovers the regulatory effects of Aβ42 on mitochondrial function, lipid metabolism, and its onco-suppressive effects accompanied by reduced synthesis of extracellular matrix proteins. Combined, these data reveal Aβ as an indispensable regulator of cell-cell interactions in healthy liver and a powerful protector against liver fibrosis.

Comparison between the Regenerative and Therapeutic Impacts of Bone Marrow Mesenchymal Stem Cells and Adipose Mesenchymal Stem Cells Pre-Treated with Melatonin on Liver Fibrosis.

The distinctive feature of liver fibrosis is the progressive replacement of healthy hepatic cells by the extracellular matrix protein, which is abundant in collagen I and III, with impaired matrix remodeling. The activation of myofibroblastic cells enhances the fibrogenic response of complex interactions of hepatic stellate cells, fibroblasts, and inflammatory cells to produce the excessive deposition of the extracellular protein matrix. This process is activated by multiple fibrogenic mediators and cytokines, such as TNF-α and IL-1β, accompanied with a decrease in the anti-fibrogenic factor NF-κβ. Mesenchymal stem cells (MSCs) represent a promising therapy for liver fibrosis, allowing for a more advanced regenerative influence when cultured with extrinsic or intrinsic proliferative factors, cytokines, antioxidants, growth factors, and hormones such as melatonin (MT). However, previous studies showed conflicting findings concerning the therapeutic effects of adipose (AD) and bone marrow (BM) MSCs; therefore, the present work aimed to conduct a comparative and comprehensive study investigating the impact of MT pre-treatment on the immunomodulatory, anti-inflammatory, and anti-apoptotic effects of AD- and BM-MSCs and to critically analyze whether MT-pre-treated AD-MSCs and BM-MSCs reveal equal or different therapeutic and regenerative potentials in a CCl4-injured liver experimental rat model. Six groups of experimental rats were used, with ten rats in each group: group I (control group), group II (CCl4-treated group), group III (CCl4- and BM-MSC-treated group), group IV (CCl4 and MT-pre-treated BM-MSC group), group V (CCl4- and AD-MSC-treated group), and group VI (CCl4 and MT-pre-treated AD-MSC group). Liver function tests and the gene expression of inflammatory, fibrogenic, apoptotic, and proliferative factors were analyzed. Histological and immunohistochemical changes were assessed. The present study compared the ability of AD- and BM-MSCs, with and without MT pre-treatment, to reduce hepatic fibrosis. Both types of MSCs improved hepatocyte function by reducing the serum levels of ALT, aspartate aminotransferase (AST), alkaline phosphatase (AKP), and total bilirubin (TBIL). In addition, the changes in the hepatocellular architecture, including the hepatocytes, liver sinusoids, central veins, portal veins, biliary ducts, and hepatic arteries, showed a decrease in hepatocyte injury and cholestasis with a reduction in inflammation, apoptosis, and necrosis of the hepatic cells, together with an inhibition of liver tissue fibrosis. These results were augmented by an analysis of the expression of the pro-inflammatory cytokines TNFα and IL-1β, the anti-fibrogenic factor NF-κβ, the apoptotic factor caspase-3, and the proliferative indicators antigen Ki-67 and proliferating cell nuclear antigen (PCNA). These findings were found to be statistically significant, with the restoration of normal parameters in the rats that received AD-MSCs pre-treated with MT, denoting optimal regenerative and therapeutic effects. AD-MSCs pre-treated with MT are the preferred choice in improving hepatic fibrosis and promoting the therapeutic and regenerative ability of liver tissue. They represent a very significant tool for future stem cell use in the tissue regeneration strategy for the treatment of liver diseases.

Liver sinusoidal cells eliminate blood-borne phage K1F.

Phage treatment has regained attention due to an increase in multiresistant bacteria. For phage therapy to be successful, phages must reach their target bacteria in sufficiently high numbers. Blood-borne phages are believed to be captured by macrophages in the liver and spleen. Since liver sinusoids also consist of specialized scavenger liver sinusoidal endothelial cells (LSECs) and Kupffer cells (KCs), this study investigated the contribution of both cell types in the elimination of Escherichia coli phage K1Fg10b::gfp (K1Fgfp) in mice. Circulatory half-life, organ, and hepatocellular distribution of K1Fgfp were determined following intravenous administration. Internalization of K1Fgfp and effects of phage opsonization on uptake were explored using primary mouse and human LSEC and KC cultures. When inoculated with 107 virions, >95% of the total K1Fgfp load was eliminated from the blood within 20 min, and 94% of the total retrieved K1Fgfp was localized to the liver. Higher doses resulted in slower elimination, possibly reflecting temporary saturation of liver scavenging capacity. Phage DNA was detected in both cell types, with a KC:LSEC ratio of 12:1 per population following cell isolation. Opsonization with plasma proteins increased time-dependent cellular uptake in both LSECs and KCs in vitro. Internalized phages were rapidly transported along the endocytic pathway to lysosomal compartments. Reduced viability of intracellular K1Fgfp corroborated inactivation following endocytosis. This study is the first to identify phage distribution in the liver at the hepatocellular level, confirming clearance of K1Fgfp performed mostly by KCs with a significant uptake also in LSECs.IMPORTANCEFaced with the increasing amounts of bacteria with multidrug antimicrobial resistance, phage therapy has regained attention as a possible treatment option. The phage field has recently experienced an emergence in commercial interest as research has identified new and more efficient ways of identifying and matching phages against resistant superbugs. Currently, phages are unapproved drugs in most parts of the world. For phages to reach broad clinical use, they must be shown to be clinically safe and useful. The results presented herein contribute to increased knowledge about the pharmacokinetics of the T7-like phage K1F in the mammalian system. The cell types of the liver that are responsible for rapid phage blood clearance are identified. Our results highlight the need for more research about appropriate dose regimens when phage therapy is delivered intravenously and advise essential knowledge about cell systems that should be investigated further for detailed phage pharmacodynamics.

SENP1 attenuates hypoxia‑reoxygenation injury in liver sinusoid endothelial cells by relying on the HIF‑1α signaling pathway.

Liver sinusoidal endothelial cells (LSECs) have an important role in hepatic ischemia‑reperfusion injury (I/R), but the specific molecular mechanism of action is unknown. LSEC proliferation is regulated and fenestration is maintained via the Sentrin/SUMO‑specific protease 1 (SENP1)/hypoxia‑inducible factor‑1α (HIF‑1α) signaling axis under hypoxic conditions. In the present study, a hypoxia‑reoxygenation (H‑R) injury model was established using mouse LSECs to explore the relationship between SENP1 and H‑R injury invitro, and the specific underlying mechanism was identified, revealing new targets for the clinical attenuation of hepatic I/R injury. Following the culture of LSECs under H‑R conditions, it was demonstrated that the expression of SENP1 was upregulated by reverse transcription‑quantitative polymerase chain reaction and western blotting (WB). In addition, scanning electron microscopy indicated that fenestrae damage was increased, a Cell Counting Kit‑8 assay demonstrated that the proliferation of cells was impaired and flow cytometry showed that apoptosis was increased. After silencing SENP1 expression with short interfering RNA, the proliferation activity of LSECs decreased, the fenestrae damage increased, the apoptosis rate increased and the expression levels of SENP1, HIF‑1α, heme oxygenase and Bcl‑2 were downregulated (as demonstrated by WB), while the expression levels of apoptosis‑related proteins, cleaved‑caspase‑3 and Bax, were upregulated. Enzyme‑linked immunosorbent assay detection showed that the level of vascular endothelial growth factor in the supernatant decreased and the level of IL‑6 and TNF‑α increased. Following the administration of an HIF‑1α signaling pathway agonist, the situation was reversed. These results therefore suggested that SENP1 attenuated the reduction in proliferation, apoptosis and fenestration of LSECs observed following H‑R injury through the HIF‑1α signaling pathway. In conclusion, SENP1 may attenuate H‑R injury in LSECs in a HIF‑1α signaling pathway‑dependent manner.

Unveiling the Therapeutic Potential of Vitamin-E in Preventing Inflammation and Stromal Congestion in Alcoholic Liver Injury.

The objective of the research was to investigate and assess how effective Vitamin E is in preventing or reducing liver inflammation and stromal congestion associated with alcoholic liver injury. This is a laboratory-based experimental study. A total of 18 domestic rabbits were divided into groups A, B, and C. Group A was the control group and treated with normal saline as a placebo. Groups B and C were given 30% ethanol in a daily dose of 30 ml/kg/day. Additionally, group C was treated with vitamin E at 50 mg/kg/day. All three groups were sub-divided into two sub-groups I and II on the basis of experimental duration of eight weeks and four weeks respectively. The subgroups with eight weeks of experimental time duration were categorized as "category E8" and subgroups with an experimental duration of four weeks were categorized as "category E4". Liver tissue samples from each animalwere subjected to staining using hematoxylin and eosin (H&E) stain for histological staining in order to assess portal inflammation and to measure the sizes of hepatic sinusoids and central veins to evaluate hepatic congestion. A statistically significant variance was observed in the size of central veins, hepatic sinusoids, and invasion of inflammatory cells in portal areas across and between the groups within categories E4 and E8. Animals treated with vitamin E exhibited lower invasion of inflammatory cells and larger central veins and sinusoids compared to those not treated with vitamin E. Vitamin E may have a significant role in reducing or limiting the infiltration of inflammatory cells and could help prevent hepatic congestion in cases of alcoholic liver injury.

Large extracellular vesicle (EV) and neutrophil extracellular trap (NET) interaction captured in vivo during systemic inflammation

Extracellular vesicles (EVs) and neutrophil extracellular traps (NETs) are pivotal bioactive structures involved in various processes including inflammation. Herein we report the interactions between EVs and NETs during murine endotoxemia studied in situ directly in the vasculature (cremaster muscle, liver sinusoids) using intravital microscopy (IVM). We captured NETs and EV release in real time by both non- and polarized neutrophils in liver but not in cremaster vasculature. When comparing numbers of circulating EVs of various origin (nanoparticle tracking analysis—NTA, flow cytometry) with those interacting with endothelium and NETs (IVM) we observed that whereas platelet and monocyte/macrophage-derived EVs dominate in blood and peritoneal lavage, respectively, mostly neutrophil-derived EVs interact with the vascular lining, NETs and leukocytes. Despite the interaction, NETs do not affect EV formation as NET release inhibition did not alter EV release. However, EVs inhibit NETs formation and in particular, erythrocyte-derived EVs downregulate NET release and this effect is mediated via Siglec-E-dependent interactions with neutrophils. Overall, we report that EVs are present in NETs in vivo and they do modulate their release but the process in not bidirectional. Moreover, EVs isolated from body fluids might not reflect their importance in direct endothelial- and leukocyte-related interactions.

Evaluation of the healing and protective properties of adipose-derived mesenchymal stem cells from cisplatin-induced liver and kidney damage.

The occurrence of nephrotoxicity and hepatotoxicity as a result of cisplatin administration is a major concern in clinical practice. This study examined the potential protective effects of administering mesenchymal stem cells (MSCs) on the renal and hepatic damage caused by cisplatin. Moreover, the study investigated the potential protective effects of administering Adipose-Derived Mesenchymal Stem Cells (ADMSC) to counteract the harmful effects of cisplatin-induced kidney and liver damage. Male Sprague-Dawley rats were divided into three groups: normal control, cisplatin + saline, and cisplatin + ADMSC. Cisplatin was administered to induce toxicity, and ADMSC was administered intravenously as a potential therapeutic intervention. Biochemical parameters and histopathological changes were assessed in the kidney and liver tissues. Statistical analyses were performed using a one-way ANOVA. Cisplatin increased malondialdehyde (MDA), tumor necrosis factor alfa (TNF-alfa), IL-6, alanine transaminase (ALT), creatinine, Galectin-3, Tissue growth factor beta 1 (TGF-beta 1), compared to the normal control group. Cisplatin-MSC reduced these levels. Histopathology showed that cisplatin caused kidney tubular epithelial necrosis, luminal necrotic debris, tubular dilatation, interstitial inflammation, liver sinusoidal and central vein dilatation, congestion, necrosis, and cytoplasmic vacuolization. ADMSC administration significantly reduced histopathological changes. These findings highlight the potential therapeutic benefits of mesenchymal stem cell (MSC) administration in mitigating cisplatin-induced nephrotoxicity and hepatotoxicity. MSC treatment demonstrated protective effects by reducing oxidative stress, inflammatory markers, and histopathological alterations. Further investigations are warranted to elucidate the precise mechanisms underlying these protective effects and evaluate their clinical implications for managing cisplatin-induced organ damage.

Abstract WP291: Cerebral Amyloid Angiopathy Induces Liver Inflammation in Aged Mice

Introduction: Cerebral amyloid angiopathy (CAA) is a chronic age-related neurodegenerative disease, identified by the presence of amyloid-beta (Aβ) deposition within the vasculature of the brain, leading to stroke and progressive cognitive decline. The liver is a key peripheral organ involved in the clearance of excess Aβ in the circulation. However, the role of the brain-liver axis in CAA is unknown. Here, we investigate inflammatory changes in the liver in a murine model of CAA. Methods: Aged (>23 months) male and female Tg-SwDI (CAA) mice harboring Swedish, Dutch, and Iowa mutations of the human amyloid precursor protein were used as a mouse model of CAA along with age-matched wild-type (WT) controls (n=3-7/grp). Hepatic inflammation was analyzed with histological and immunofluorescence imaging. Single cells from liver tissue were isolated and analyzed using flow cytometry. Total RNA was isolated to examine gene expression in the liver using real-time quantitative PCR. Results: Our data showed that CAA increases infiltration of immune cells proximal to liver sinusoids across both sexes compared to WT mice. We found increased macrophages (i.e., F4/80 + cells) in the liver of CAA mice compared to WT controls. Flow cytometry on the liver revealed that female CAA mice had a significant increase in dendritic cells (i.e., CD80 + CD45 + CD11c + cells, *** P <0.001) compared to WT females. Further, we found that Oatp2 , a membrane transporter which mediates hepatocytic uptake of toxic molecules (e.g., Aβ), is associated with sex-specific CAA pathophysiology in the liver. Conclusions: In this study, we found that sex differences may exist in CAA-induced liver inflammation. Further investigations are needed to study how vascular amyloidosis shapes the immune responses and peripheral Aβ clearance in the context of aging and CAA.

Portal hypertension in patients with nonalcoholic fatty liver disease: Current knowledge and challenges.

Portal hypertension (PH) has traditionally been observed as a consequence of significant fibrosis and cirrhosis in advanced non-alcoholic fatty liver disease (NAFLD). However, recent studies have provided evidence that PH may develop in earlier stages of NAFLD, suggesting that there are additional pathogenetic mechanisms at work in addition to liver fibrosis. The early development of PH in NAFLD is associated with hepatocellular lipid accumulation and ballooning, leading to the compression of liver sinusoids. External compression and intra-luminal obstacles cause mechanical forces such as strain, shear stress and elevated hydrostatic pressure that in turn activate mechanotransduction pathways, resulting in endothelial dysfunction and the development of fibrosis. The spatial distribution of histological and functional changes in the periportal and perisinusoidal areas of the liver lobule are considered responsible for the pre-sinusoidal component of PH in patients with NAFLD. Thus, current diagnostic methods such as hepatic venous pressure gradient (HVPG) measurement tend to underestimate portal pressure (PP) in NAFLD patients, who might decompensate below the HVPG threshold of 10 mmHg, which is traditionally considered the most relevant indicator of clinically significant portal hypertension (CSPH). This creates further challenges in finding a reliable diagnostic method to stratify the prognostic risk in this population of patients. In theory, the measurement of the portal pressure gradient guided by endoscopic ultrasound might overcome the limitations of HVPG measurement by avoiding the influence of the pre-sinusoidal component, but more investigations are needed to test its clinical utility for this indication. Liver and spleen stiffness measurement in combination with platelet count is currently the best-validated non-invasive approach for diagnosing CSPH and varices needing treatment. Lifestyle change remains the cornerstone of the treatment of PH in NAFLD, together with correcting the components of metabolic syndrome, using nonselective beta blockers, whereas emerging candidate drugs require more robust confirmation from clinical trials.

Pathological effects of pregabalin toxicity in rats

Abstract Background: Pregabalin (PGB) is one kind of gabapentinoid. The main mechanism of action is binding at the alpha-2-delta site, which inhibits calcium influx in response to depolarization at nerve terminals and, in turn, suppresses the release of glutamate, noradrenaline, and substance P. Objectives: This study aimed to study the pathological effects of PGB toxicity on brain and liver of rats. Materials and Methods: We chose 20 mature albino rats, both sexes, averaging 220 g in weight; these were divided into two groups (10 rats per group): the toxic group and the control group. Tablets of Lyrica (Pfizer) may be purchased over the counter. The 300 mg of PGB in each tablet was dissolved in 3 mL of 0.9% normal saline. The dosage was determined using the maximum lethal oral dose in rats (5000 mg/kg) (Pfizer, 2017). Based on the rats’ weights, a toxic dosage of 1000 mg of PGB was determined and reconstituted in normal saline (0.9% in 3 mL). Each animal in the acute toxicity group received a single dose of the produced medication orally. After 24 h, all of the animals in both groups were euthanized. The brain and liver were quickly dissected and removed, washed in saline solution, and then processed for histopathological study. Results: Focal regions of hemorrhage and congestion were seen in H&E-stained sections from the acute toxicity group, and most pyramidal cells were degraded, pyknotic, and exhibited karyolysis. Cerebellar cortical layers were preserved; however, Purkinje cells were destroyed in the acute toxicity group, which also exhibited an increase in pyknotic cells, hemorrhage, vascular congestion, and localized loss of tissue. Hemorrhages, congestion of portal region blood vessels, and central veins and hepatic sinusoids were some of the most notable pathological abnormalities seen in the livers of those using PGB. Hepatocytes show nuclear pyknosis and a homogeneously acidophilic cytoplasm as a result of severe degenerative alterations, such as vacuolar degeneration and severe necrotic changes. Conclusion: PGB can cause pathological lesions in the brain and liver at a single toxic dose.

Exposure to tobacco (Nicotiana tabacum) extract and cigarette smoke induces hematological and histopathological alterations in Swiss albino mice

Tobacco smoking is a widely practiced recreational activity and is associated with numerous health risks. Hence, the present study was conducted on Swiss albino mice to assess the impact of aqueous extract of tobacco and cigarette smoke on behavioral, hematological, and histopathological parameters of the lungs, liver, and kidney. The mice were divided into Control (Cont), Tobacco nicotiana (TN), and Cigarette smoke (CS) groups (n=6). TN group received aqueous tobacco orally at a dose of 5ml/kg BW for 21 days, whereas CS group was exposed to inhalational commercial cigarette smoke for the same duration. On day 22, blood samples and visceral organs were collected to evaluate hematological (TEC, TLC, Hb%) and histopathological parameters from euthanized mice. The TEC values were significantly elevated (P<0.05) in both groups. In contrast to the control group, the TLC value was lower in the CS group (P>0.05) than in the TN group, but the mean Hb% values showed no significant changes. Lower body weight, aggressive behavior, and weakness were observed in the treated groups. Histopathologically, the lung revealed emphysematous changes, alveolar septal thickening, inflammatory cells infiltration, congestion, and thrombus formation in pulmonary artery. Liver exhibited hepatocyte degeneration with karyomegaly and karyorrhexis, derangement of hepatic cords and sinusoids with central vein congestion. Derangement of renal tubules, edema, and glomerular degeneration observed in the kidney. These observations lead us to postulate that tobacco and cigarette smoke exert detrimental effects on the structure of some vital organs in the body after direct or indirect uptake.

3D bioprinting of Gelatin-Alginate bioinks for biofabrication of in vitro liver sinusoid model

In vitro liver models are pivotal in early drug development as they serve in assessing drug toxicity. Currently, most of the liver models include static cultures of liver derived cell spheroids. Mimicking dynamic conditions (similar to native liver – such as blood flow, glucose gradients etc.) in in vitro liver models would be essential for studying drug induced liver injury (DILI). The current work investigated and developed a simple bioink composed of gelatin and alginate blends (GA) that can be used to 3D bioprint hepatocyte laden scaffolds, serving as an in vitro liver model for drug toxicity assays.Rheological evaluation (using a rotary rheometer) of various compositions of GA blends was performed to assess their printability with an extrusion bioprinter. Suitable GA compositions with human liver carcinoma cells (HepG2) were 3D bioprinted. The scaffolds were stabilized by crosslinking the alginate (using calcium chloride) and gelatin (using microbial transglutaminase). Cell viability and proliferation of HepG2 in bioprinted scaffolds was assessed for 28 days. Further work includes fabrication of HepG2 laden hollow tubes and colonization of inner lumen with human umbilical vein endothelial cells (HUVEC), followed by DILI studies in long-term perfusion culture.

Hepatic venous gas secondary to pulmonary barotrauma: rat model study.

Intrahepatic gas (IHG) is commonly observed during early postmortem examinations of humans with upper or lower airway obstructions. We conducted a study to test the hypothesis that intrapulmonary gas could retrogradely spread to the hepatic vein following pulmonary barotrauma (PB). To establish a rat model of pulmonary barotrauma, we utilized a controllable pressure-vacuum pump to apply airway pressure (40, 60, or 80mmHg). The rats were dissected directly at the end of the experiment, and histological analysis was performed through microscopic examination of the rats. Additionally, the rats were ventilated with meglumine diatrizoate under pressures of 160 and 250mmHg to observe the signal dynamic diffusion using X-ray fluoroscopy examination. Rats exhibited classical changes associated with PB, such as alveolar rupture, pulmonary interstitial emphysema, and hemorrhage, as well as IHG characterized by the presence of gas in the hepatic vein and hepatic sinusoids. Air emboli were not observed in the liver in any of the 40mmHg groups. However, they were observed in the liver in the 60 and 80mmHg groups,the amount and size of air emboli in the 80mmHg group were greater than those in the 60mmHg group (p < 0.05). The 80mmHg group presented radial grape-like bubbles in the centrilobular portion of the liver accompanied by congestion in the peripheral region of the hepatic lobule. X-ray fluoroscopy examination revealed a gradual enhancement of dynamic contrast medium signals from the lung to the inferior vena cava and then to the liver. Our findings indicate that pulmonary barotrauma can lead to the retrograde spread of intrapulmonary gas to the hepatic vein. When it is clear that no decomposition of the body has occurred, the presence of IHG serves as a novel indicator for the diagnosis of obstructive pulmonary disease or obstruction in the upper or lower airway.

Study of the f-cell ratio using plasma dilution and albumin mass kinetics

BackgroundThe f-cell ratio of 0.91 is a conversion factor between the hematocrit measured in peripheral blood and the hematocrit obtained by separate measurements of the red blood cell mass and plasma volume. The physiological background of the f-cell ratio is unclear. MethodsData were retrieved from 155 intravenous infusion experiments where 15–25 mL/kg of crystalloid fluid diluted the blood hemoglobin and plasma albumin concentrations. The hemodilution was converted to plasma dilution using the peripheral hematocrit, and the volume of distribution of exogenous albumin was calculated in 41 volunteers who received 20 % or 5 % albumin by intravenous infusion. Finally, the kinetics of plasma albumin was studied during 98 infusion experiments with 20 % albumin. ResultsPlasma dilution based on hemoglobin and albumin showed a median difference of −0.001 and a mean difference of 0.000 (N = 2184), which demonstrates that these biomarkers indicate the same expandable vascular space. In contrast, exogenous albumin occupied a volume that was 10 % larger than the plasma volume indicated by the anthropometric equations of Nadler et al. and Retzlaff et al. The kinetic analysis identified a secondary compartment that was 450 mL in size and rapidly exchanged albumin with the circulating plasma. ConclusionsThe results suggest that the f-cell ratio is due to rapid exchange of albumin between the plasma and a non-expandable compartment located outside the circulating blood (possibly the liver sinusoids). This means that the hematocrit measured in peripheral blood correctly represents the ratio between the red cell volume and the circulating plasma volume.

Endoglin and soluble endoglin in liver sinusoidal endothelial dysfunction in vivo

Liver sinusoidal endothelial cells (LSECs) play a crucial role in regulating the hepatic function. Endoglin (ENG), a transmembrane glycoprotein, was shown to be related to the development of endothelial dysfunction. In this study, we hypothesized the relationship between changes in ENG expression and markers of liver sinusoidal endothelial dysfunction (LSED) during liver impairment.Male C57BL/6J mice aged 9–12 weeks were fed with 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC) diet (intrahepatic cholestasis) or choline-deficient l-amino acid defined high-fat diet (CDAA-HFD) (non-alcoholic steatohepatitis (NASH)).Significant increases in liver enzymes, fibrosis, and inflammation biomarkers were observed in both cholestasis and NASH. Decreased p-eNOS/eNOS and VE-cadherin protein expression and a significant increase in VCAM-1 and ICAM-1 expression were detected, indicating LSED in both mouse models of liver damage. A significant reduction of ENG in the DDC-fed mice, while a significant increase of ENG in the CDAA-HFD group was observed. Both DDC and CDAA-HFD-fed mice showed a significant increase in MMP-14 protein expression, which is related to significantly increased levels of soluble endoglin (sENG) in the plasma.In conclusion, we demonstrated that intrahepatic cholestasis and NASH result in an altered ENG expression, predominantly in LSECs, suggesting a critical role of ENG expression for the proper function of liver sinusoids. Both pathologies resulted in elevated sENG levels, cleaved by MMP-14 expressed predominantly from LSECs, indicating sENG as a liver injury biomarker.