Latent Diffusion Models with Image-Derived Annotations for Enhanced AI-Assisted Cancer Diagnosis in Histopathology.

Artificial Intelligence (AI)-based image analysis has immense potential to support diagnostic histopathology, including cancer diagnostics. However, developing supervised AI methods requires large-scale annotated datasets. A potentially powerful solution is to augment training data with synthetic data. Latent diffusion models, which can generate high-quality, diverse synthetic images, are promising. However, the most common implementations rely on detailed textual descriptions, which are not generally available in this domain. This work proposes a method that constructs structured textual prompts from automatically extracted image features. We experiment with the PCam dataset, composed of tissue patches only loosely annotated as healthy or cancerous. We show that including image-derived features in the prompt, as opposed to only healthy and cancerous labels, improves the Fréchet Inception Distance (FID) by 88.6. We also show that pathologists find it challenging to detect synthetic images, with a median sensitivity/specificity of 0.55/0.55. Finally, we show that synthetic data effectively train AI models.

Runcaciguat, a novel soluble guanylate cyclase activator, shows renoprotection in hypertensive, diabetic, and metabolic preclinical models of chronic kidney disease.

Chronic kidney diseaQueryse (CKD) is associated with oxidative stress which can interrupt the nitric oxide (NO)/soluble guanylyl cyclase (sGC) signaling and decrease cyclic guanosine monophosphate (cGMP) production. Low cGMP concentrations can cause kidney damage and progression of CKD. The novel sGC activator runcaciguat targets the oxidized and heme-free form of sGC, restoring cGMP production under oxidative stress. The purpose of this study is to investigate if runcaciguat could provide an effective treatment for CKD. Runcaciguat was used for the treatment not only in rat CKD models with different etiologies and comorbidities, namely of hypertensive rats, the renin transgenic (RenTG) rat, and angiotensin-supplemented (ANG-SD) rat, but also in rats with diabetic and metabolic CKD, the Zucker diabetic fatty (ZDF) rat. The treatment duration was 2 to 42 weeks and runcaciguat was applied orally in doses from 1 to 10 mg/kg/bid. In these different rat CKD models, runcaciguat significantly reduced proteinuria (urinary protein to creatinine ratio; uPCR). These effects were also significant at doses which did not or only moderately decrease systemic blood pressure. Moreover, runcaciguat significantly decreased kidney injury biomarkers and attenuated morphological kidney damages. In RenTG rats, runcaciguat improved survival rates and markers of heart injury. These data demonstrate that the sGC activator runcaciguat exhibits cardio-renal protection at doses which did not reduce blood pressure and was effective in hypertensive as well as diabetic and metabolic CKD models. These data, therefore, suggest that runcaciguat, with its specific mode of action, represents an efficient treatment approach for CKD and associated CV diseases.

Artificial Intelligence in Toxicologic Pathology: Quantitative Evaluation of Compound-Induced Hepatocellular Hypertrophy in Rats.

Digital pathology evolved rapidly, enabling more systematic usage of image analysis and development of artificial intelligence (AI) applications. Here, combined AI models were developed to evaluate hepatocellular hypertrophy in rat liver, using commercial AI-based software on hematoxylin and eosin-stained whole slide images. In a first approach, deep learning-based identification of critical tissue zones (centrilobular, midzonal, and periportal) enabled evaluation of region-specific cell size. Mean cytoplasmic area of hepatocytes was calculated via several sequential algorithms including segmentation in microanatomical structures (separation of sinusoids and vessels from hepatocytes), nuclear detection, and area measurements. An increase in mean cytoplasmic area could be shown in groups given phenobarbital, known to induce hepatocellular hypertrophy when compared to control groups, in multiple studies. Quantitative results correlated with the gold standard: observation and grading performed by board-certified veterinary pathologists, liver weights, and gene expression. Furthermore, as a second approach, we introduce for the first time deep learning-based direct detection of hepatocellular hypertrophy with similar results. Cell hypertrophy is challenging to pick up, particularly in milder cases. Additional evaluation of mean cytoplasmic area or direct detection of hypertrophy, combined with histopathological observations and liver weights, is expected to increase accuracy and repeatability of diagnoses and grading by pathologists.

Energy metabolism modulation by biguanides in comparison with rotenone in rat liver and heart.

The biguanide metformin, a widely used antidiabetic drug, has received great interest in oncology research in recent years after an epidemiological study showed a link between metformin treatment and a reduced cancer risk in diabetic patients. Since mitochondrial metabolism has become a target for possible cancer therapeutic approaches, especially for tumors relying on oxidative metabolism, mitochondrial complex I inhibition is under discussion to be responsible for the anti-cancer effect of metformin. Rotenone, a well-known strong mitochondrial complex I inhibitor, yet associated with toxic effects, has also shown anti-cancer activity. Thus, we compared metformin and phenformin, another biguanide previously on the market as antidiabetic, with rotenone, to elucidate potential mechanisms rendering biguanides apparently less toxic than rotenone. Therefore, we conducted in vivo rat studies with metformin and phenformin, based on an experimental design previously described for mechanistic investigations of the effects of rotenone, including blood and tissue analysis, histopathology and gene expression profiling. These investigations show that the mechanistic profile of phenformin appears similar to that of rotenone, yet at a quantitatively reduced level, whereas metformin displays only transient similarities after one day of treatment. A potential reason may be that metformin, but not rotenone or phenformin, self-limits its entry into mitochondria due to its molecular properties. Thus, our detailed molecular characterization of these compounds suggests that inhibition of mitochondrial functions can serve as target for an anti-cancer mode of action, but should be self-limited or balanced to some extent to avoid exhaustion of all energy stores.

Mechanistic Investigations of the Mitochondrial Complex I Inhibitor Rotenone in the Context of Pharmacological and Safety Evaluation.

Inhibitors of the mitochondrial respiratory chain complex I are suggested to exert anti-tumor activity on those tumors relying on oxidative metabolism and are therefore of interest to oncology research. Nevertheless, the safety profile of these inhibitors should be thoroughly assessed. Rotenone, a proven complex I inhibitor, has shown anti-carcinogenic activity in several studies. In this context rotenone was used in this study as a tool compound with the aim to identify suitable biomarker candidates and provide enhanced mechanistic insights into the molecular and cellular effects of complex I inhibitors. Rats were treated with 400 ppm rotenone daily for 1, 3 or 14 consecutive days followed by necropsy. Classical clinical endpoints, including hematology, clinical chemistry and histopathology with supporting investigations (FACS-analysis, enzymatic activity assays) were examined as well as gene expression analysis. Through these investigations, we identified liver, bone marrow and bone as target organs amongst approx. 40 organs evaluated at least histopathologically. Our results suggest blood analysis, bone marrow parameters, assessment of lactate in serum and glycogen in liver, and especially gene expression analysis in liver as useful parameters for an experimental model to help to characterize the profile of complex I inhibitors with respect to a tolerable risk-benefit balance.

Chronic Activation of Heme Free Guanylate Cyclase Leads to Renal Protection in Dahl Salt-Sensitive Rats.

The nitric oxide (NO)/soluble guanylate cyclase (sGC)/cyclic guanosine monophasphate (cGMP)-signalling pathway is impaired under oxidative stress conditions due to oxidation and subsequent loss of the prosthetic sGC heme group as observed in particular in chronic renal failure. Thus, the pool of heme free sGC is increased under pathological conditions. sGC activators such as cinaciguat selectively activate the heme free form of sGC and target the disease associated enzyme. In this study, a therapeutic effect of long-term activation of heme free sGC by the sGC activator cinaciguat was investigated in an experimental model of salt-sensitive hypertension, a condition that is associated with increased oxidative stress, heme loss from sGC and development of chronic renal failure. For that purpose Dahl/ss rats, which develop severe hypertension upon high salt intake, were fed a high salt diet (8% NaCl) containing either placebo or cinaciguat for 21 weeks. Cinaciguat markedly improved survival and ameliorated the salt-induced increase in blood pressure upon treatment with cinaciguat compared to placebo. Renal function was significantly improved in the cinaciguat group compared to the placebo group as indicated by a significantly improved glomerular filtration rate and reduced urinary protein excretion. This was due to anti-fibrotic and anti-inflammatory effects of the cinaciguat treatment. Taken together, this is the first study showing that long-term activation of heme free sGC leads to renal protection in an experimental model of hypertension and chronic kidney disease. These results underline the promising potential of cinaciguat to treat renal diseases by targeting the disease associated heme free form of sGC.

Evaluation of the Adulticidal Efficacy of Imidacloprid 10 %/Moxidectin 2.5 % (w/v) Spot-on (Advocate®, Advantage® Multi) against Dirofilaria repens in Experimentally Infected Dogs.

This study aimed to evaluate the efficacy of imidacloprid 10 %/moxidectin 2.5 % (w/v) spot-on (Advocate®/Advantage® Multi, Bayer) against adult Dirofilaria repens in a blinded, placebo-controlled randomised laboratory study. Twenty-four Beagle dogs were experimentally infected with approximately 75 infective D. repens larvae each on study day (SD) 0. Treatment was initiated on SD 228 after patency had been confirmed in 21 dogs, using a modified Knott Test. Eleven dogs received monthly treatments with imidacloprid/moxidectin at the minimum therapeutic dose (10 mg/kg imidacloprid and 2.5 mg/kg moxidectin) for six consecutive months and 12 control dogs were treated with a placebo formulation. Approximately one month after the last treatment, all dogs were euthanised and necropsied for the detection of D. repens worms. Eleven control dogs harboured live adult D. repens (range 2-11, geometric mean 5.44). Eight of 11 imidacloprid/moxidectin-treated dogs were free of live worms. The live worm count was reduced by 96.2 % (range 0-1, geometric mean 0.21). The majority of dead worms were encapsulated and degenerated. After the first treatment, Knott Tests were negative in all imidacloprid/moxidectin-treated dogs and this status was maintained in 10 dogs until study end. One dog showed a low microfilariae count (1 and 4/mL) on four occasions but was also negative before necropsy. The treatment was well tolerated by all study animals. It is concluded that six consecutive monthly treatments with imidacloprid/moxidectin spot-on are effective and safe against adult D. repens and provide an option for preventing the further spread of this zoonotic parasite.

Soluble guanylate cyclase stimulation prevents fibrotic tissue remodeling and improves survival in salt-sensitive Dahl rats.

BACKGROUND: A direct pharmacological stimulation of soluble guanylate cyclase (sGC) is an emerging therapeutic approach to the management of various cardiovascular disorders associated with endothelial dysfunction. Novel sGC stimulators, including riociguat (BAY 63-2521), have a dual mode of action: They sensitize sGC to endogenously produced nitric oxide (NO) and also directly stimulate sGC independently of NO. Little is known about their effects on tissue remodeling and degeneration and survival in experimental malignant hypertension. METHODS AND RESULTS: Mortality, hemodynamics and biomarkers of tissue remodeling and degeneration were assessed in Dahl salt-sensitive rats maintained on a high salt diet and treated with riociguat (3 or 10 mg/kg/d) for 14 weeks. Riociguat markedly attenuated systemic hypertension, improved systolic heart function and increased survival from 33% to 85%. Histological examination of the heart and kidneys revealed that riociguat significantly ameliorated fibrotic tissue remodeling and degeneration. Correspondingly, mRNA expression of the pro-fibrotic biomarkers osteopontin (OPN), tissue inhibitor of matrix metalloproteinase-1 (TIMP-1) and plasminogen activator inhibitor-1 (PAI-1) in the myocardium and the renal cortex was attenuated by riociguat. In addition, riociguat reduced plasma and urinary levels of OPN, TIMP-1, and PAI-1. CONCLUSIONS: Stimulation of sGC by riociguat markedly improves survival and attenuates systemic hypertension and systolic dysfunction, as well as fibrotic tissue remodeling in the myocardium and the renal cortex in a rodent model of pressure and volume overload. These findings suggest a therapeutic potential of sGC stimulators in diseases associated with impaired cardiovascular and renal functions.

Nitric oxide-independent stimulation of soluble guanylate cyclase reduces organ damage in experimental low-renin and high-renin models.

OBJECTIVES: The nitric oxide-soluble guanylate cyclase (sGC)-cGMP signal transduction pathway is impaired in different cardiovascular diseases, including pulmonary hypertension, heart failure and arterial hypertension. Riociguat is a novel stimulator of soluble guanylate cyclase (sGC). However, little is known about the effects of sGC stimulators in experimental models of hypertension. We thus investigated the cardio-renal protective effects of riociguat in low-renin and high-renin rat models of hypertension. METHODS: The vasorelaxant effect of riociguat was tested in vitro on isolated saphenous artery rings of normal and nitrate tolerant rabbits. The cardiovascular in-vivo effects of sGC stimulation were evaluated in hypertensive renin-transgenic rats treated with the nitric oxide-synthase inhibitor N-nitro-L-arginine methyl ester (L-NAME) (high-renin model) and in rats with 5/6 nephrectomy (low-renin model). RESULTS: In both animal models, riociguat treatment improved survival and normalized blood pressure. Moreover, in the L-NAME study part, riociguat reduced cardiac target organ damage as indicated by lower plasma ANP, lower relative left ventricular weight and lower cardiac interstitial fibrosis, and reduced renal target organ damage as indicated by lower plasma creatinine and urea, less glomerulosclerosis and less renal interstitial fibrosis. In the 5/6 nephrectomy study part, riociguat reduced cardiac target organ damage as indicated by lower plasma ANP, lower relative left ventricular weight, lower myocyte diameter and lower arterial media/lumen ratio, and reduced renal target organ damage as indicated by improved creatinine clearance and less renal interstitial fibrosis. CONCLUSION: We demonstrate for the first time that the novel sGC stimulator riociguat shows in two independent models of hypertension a potent protection against cardiac and renal target organ damage.