The main outcome measures were: Ocular Surface Disease Index (OSDI), Best Corrected Visual Acuity (BCVA), the Schirmer test, tear break-up time (TBUT), corneal and conjunctival staining according to the Oxford scale, conjunctival hyperaemia, and Meibomian gland parameters. The results were compared at baseline, 1 month, and 3 months following the treatment. The clinical results were correlated with the concentration of GFs in the biological tear substitutes. Results: Significant differences were observed in the concentration of FGF , EGF , fibronectin , VEGF , PDGF AB , NGF , PDGF , substance p  in PRP compared to AS. The level of TGF-β was undoubtedly higher in AS than in PRP . No significant differences between AS and PRP were observed in the concentration of IGF. Therapy with blood products relieved the signs and symptoms in pSS DE patients. There was a statistically significant improvement in BCVA, the Schirmer test, TBUT, Meibomian gland parameters, and the reduction of the OSDI scores, Oxford staining, and conjunctiva hyperaemia in each of the groups. However, the clinical changes were more significant in the PRP group. There were numerous correlations between the level of GFs and the mean change in clinical outcomes. No adverse events were reported. Conclusions: Despite the fact that blood derivatives differ in composition, they seem to be effective and safe in the treatment of severe DE in pSS patients. The signs and symptoms of DE were reduced in both groups, but only the mean change in OSDI was statistically significant. A greater reduction in OSDI scores was observed in the PRP group. The obtained results and the composition of haemoderivates may indicate the superiority of PRP in relieving the symptoms of DE in pSS patients compared to AS.Purpose: The aim of the study was to compare the difference in composition between 100% autologous serum (AS) and 100% platelet-rich plasma (PRP) eye drops and assess their impact on the clinical outcomes after the treatment of severe dry eye (DE) in primary Sjogren Syndrome patients (pSS). Materials and Methods: This is an interventional, non-randomized, comparative, three-month study. 22 patients with severe DE in pSS were treated with 100% AS (22 eyes) and 100% PRP (22 eyes) eye drops 5 times per day in monotherapy mode. The quantifications of growth factors (GFs) such as fibroblast growth factor (FGF), epidermal growth factor (EGF), vascular endothelial growth factor (VEGF), platelet-derived growth factor (PDGF), nerve growth factor (NGF), transforming growth factor (TGF-b), insulin-like growth factor (IGF), fibronectin, and substance  Sjogren’s syndrome (SS) is a complex autoimmune disease with multiple lesions resulting. The main manifestations include mouth and eye dryness, accompanied by systemic complications  . KJ BlocOcular or oral dryness is present in approximately 95–98% of pSS patients, and as many as 89% of patients present with both . Ocular Due to the multifactorial etiology, the treatment poses a challenge and presents limited efficacy. Current management of Sjogren syndrome dry eye (SSDE) includes topical administration of artificial tear substitutes such as drops, gels, or ointments whose task is lubrication of the ocular surface. Unfortunately, the substitutes lack ingredients found in natural tears, e.g., proteins, lipids, and growth factors. Moreover, in moderate or severe DE, only preservative-free artificial tears should be considered due to the harmful, toxic, or allergic effects of the common-use preservatives on the already damaged ocular surface. Considering the accompanying inflammation process, other therapeutic options are glucocorticoids and cyclosporine A, but their side effects  should not be ignored. The next step of the DE treatment is interventional procedures such as punctal occlusion, amniotic membrane transplantation, or the application of neurostimulators. Due to the limited success of artificial tears  , and platelet lysate.Autologous serum was first introduced in 1975 to treat ocular surface disease . Later, PRP is an autologous blood product with platelet concentrations above baseline . The firThe role of AS and PRP in ocular surface damage has been an area of interest for numerous ophthalmologists. The purpose of this study was to determine the differences in the composition of 100% AS and 100% PRP eye drops and to determine the impact of growth factor levels on clinical outcomes in the treatment of severe DE in pSS.This is an interventional, non-randomized, comparative study with patients recruited from the ophthalmology clinic of the Medical University of Lublin, Poland. The research was approved by the Local Ethics Committee of the Medical University of Lublin (no. KE-0254/61) and followed the tenets of the Declaration of Helsinki.Twenty-two female patients with a diagnosis of pSS in accordance with the 2002 American-European Consensus Group Criteria for Sjögren’s Syndrome (AECG)  took parWomen with dry eye disease due to primary Sjogren syndrome diagnosed according to the AECG 2002 classification ;age ≥ 18 years old;no general disease;the same diet habits ;Ocular Surface Disease Index questionnaire (OSDI ≥ 23), at least moderate DE symptoms;tear break-up time (TBUT < 10 s);Schirmer’s test (≤10 mm);abnormal ocular surface staining—Oxford Scale ≥ 2.Inclusion criteria:pregnancy or breastfeeding;other connective tissue disorders;diabetes;acute or chronic general infection, infectious disease;cancer;blood pathology or blood-transmitted disease;chronic liver or kidney disease;anticoagulant or antiplatelet therapy;use of haemoderivates within 3 months before the enrolment;glaucoma;previous history of ocular trauma, ocular surgery within 6 months;use of contact lenses;punctal plug.Exclusion criteria:Previously topically used medications (drops) were stopped 48 h before starting the AS or PRP therapy. A comprehensive ophthalmological examination, medical history, demographic data, and ocular evaluation were all recorded.The patients who met the inclusion criteria and consented to participate in the study were divided into 2 groups and advised to use the derivatives  1 drop 5 times per day. Participants were assessed by a masked research ophthalmologist (not participating in the therapeutic process) at baseline, 1 month, and 3 months into the treatment. The OSDI questionnaire was filled out. The clinical examination was performed by a trained masked ophthalmologist and included a visual acuity measurement and a comprehensive anterior segment evaluation with slit lamp biomicroscopy. Tests of tear function, such as the Schirmer test and tear breakup time, were performed, as well as an evaluation of the cornea and conjunctiva surfaces with ocular surface staining in agreement with the Oxford scale, conjunctival hyperaemia, and the Meibomian gland parameters. The tests were performed as described below before the treatment and at a 1-month and 3-month follow-up after the treatment.The OSDI standard self-assessment questionnaire consisted of 12 validated questions divided into 3 parts devoted to visual-related function, ocular symptoms, and environment-triggered symptoms . Each quSnellen VA testing was performed using commercial Snellen projectors according to a routine, standard-of-care examination, and a score was given on the basis of a line assignment method using the last line where at least 3 of 5 letters were correctly identified. Then, VA was converted to a logMAR scale according to the pattern: logMAR = −1 * log10 (snellen_frac).A Schirmer strip (Whatman filter paper number 41) was placed in the lateral third of the inferior conjunctival fornix. Patients were asked to close their eyes. The amount of wetting on the filter paper was recorded after 5 min.Moreover, unstimulated salivary Schirmer tests were performed in the morning to measure unstimulated salivary flow. Patients were not allowed to eat, drink, smoke, or brush their teeth for 2 h before the examination. Participants were asked to sit upright in a chair, swallow all the saliva in the mouth before the test, and not swallow during the test. Moreover, patients were requested to open their mouths and rest their tongues on the hard palate so as to prevent the inadvertent wetting of the test strips. The results were recorded after 5 min . The tesA fluorescein strip with alcaine  was placed at the inferior conjunctival fornix without touching the cornea. Patients were instructed to blink a few times (3–5) and to abstain from blinking during the examination. The measurements were taken during a slit lamp examination with cobalt blue light. The time taken for the first dry spot to appear after blinking was recorded, then three measurements were taken, and the average was calculated and used for analysis.The Oxford scale for corneal and conjunctival fluorescein staining patterns was graded from zero to five according to previously established criteria .The level of bulbar hyperaemia was evaluated using the Brien Holden Vision Institute scales, which included a 5-point bulbar redness photographic scale where 0 meant no hyperaemia, 1 = very slight, 2 = slight, 3 = moderate, and 4 = severe .The quality of meibomian gland secretion was assessed during a slit lamp examination in each of the eight glands of the central third of the lower lid on a scale of 0 to 3 for each gland: 0—clear, 1—cloudy, 2—cloudy with debris (granular), and 3—thick like toothpaste .Expressibility was determined on a scale of 0 to 3 in five glands in the lower or upper lid, according to the number of glands expressible: 0—all glands, 1—three to four glands, 2—one to two glands, and 3—no glands .Patients were asked about any adverse effects of the topical treatment at the 1-month and 3-month follow-up visits.AS and PRP preparation were performed under strict sterility conditions using a laminar flow hood on all patients. Patients fasted before blood sampling, which took place in the morning between 7 and 9 o’clock. Initially, 5 mL of peripheral whole blood was collected by venipuncture in the arm using EDTA tubes  to evaluate the number of platelets (PLT) and white and red blood cells.The blood was collected into 5 mL vacuum probes without anticoagulant or clot activator . The samples were left standing in an upright position for 1 h at room temperature (22–26 °C) to ensure blood clot formation. The tubes were centrifuged for 10 min at 3000 rpm to allow the separation of serum from the clotted blood. Then the supernatant serum was transferred into new tubes and centrifuged for 10 min at 3000 rpm. The serum was introduced into a Millex filtration unit  and dispensed without dilution into 2 mL Eppendorf tubes for further analysis and as the final product to be administered by the participants in the form of eye drops.Peripheral blood was collected in 9-mL vacuum tubes containing 1 mL of 3.8% sodium citrate as an anticoagulant . PRP was prepared in a single-step technique and centrifuged at 1400 rpm for 10 min according to the protocol previously described by Alió. This resulted in the formation of three layers: mostly red blood cells at the bottom, a thin middle buffer layer of leukocytes, and platelet plasma at the top of the tube. Aggregation or activation of platelets were not induced. All PRP top layers were collected, avoiding the buffy coat, and divided into smaller portions (2 mL Eppendorf tubes) for further analysis and as the final product to be administered by the participants as eye drops.Manipulation of the blood, autologous serum, and platelet-rich plasma was performed under strict sterility conditions inside the laminar flow cabin with sterile disposable materials. The blood was collected from all patients on the same day. Moreover, autologous serum and platelet-rich plasma were prepared immediately and divided into 2 mL Eppendorf tubes for further analysis. The time between blood collection, preparation of blood products, and their transfer to the laboratory did not exceed 4 h. During that time, the samples were stored at 4 °C. The transport to the laboratory was carried out in a transport refrigerator that maintains a temperature of 4 °C. Morphology determinations and ELISA tests were performed immediately after the samples delivery to the laboratory. The final blood products, AS and PRP, were transferred into opaque eye drop bottles to protect them from ultraviolet light and labeled with a name and a date. Each patient received 12 bottles for the whole treatment period and a leaflet with instructions on how to apply and store the product. The currently used bottle should have been stored at 4 °C for 7 days (one bottle for one week) and the remaining bottles at −20 °C in a fridge until the day of use.The evaluation of the total amount of blood cells  in whole blood, AS, and PRP was performed twice for each sample using automated haemocytometry . The means of the two measurements were used for further calculations.p were used to quantify their amount in AS and PRP on the day of blood collection and haemoderivate preparations. The molecules were chosen according to their functions in the corneal regeneration process. The procedures were carried out following the manufacturer’s instructions on the attached datasheet for the EuroImmune Analyzer. All samples were measured twice, and the means of the two measurements were taken into consideration in further investigations. Results were expressed as pg/mL.A commercially available enzyme-linked immunosorbent assay (ELISA) kit  specific for GFs and protein levels such as fibroblast growth factor (FGF), epidermal growth factor (EGF), vascular endothelial growth factor (VEGF), platelet-derived growth factor (PDGF), nerve growth factor (NGF), transforming growth factor (TGF-b), insulin-like growth factor (IGF), fibronectin, and substance A summary of the design of the study is presented in The statistical power was assumed at 80%, alpha < 0.05, and a relative difference of at least 30% when calculating the sample size for the study. The investigators were required to test at least 44 samples in the autologous serum arm vs. 44 in the platelet-rich plasma arm. The authors adhered to the adopted study plan and the aforementioned sample sizes. Each patient provided two test tubes, i.e., one containing AS and the other containing PRP. Subsequently, each sample vial was subjected to laboratory measurements twice.Categorical variables were described using integer numbers and percentages. Numerical traits were depicted through their mean, standard deviation, median, lower-to-upper quartile, and minimum-to-maximum values.The normality of the distribution was assessed using the Shapiro-Wilk W test. The homogeneity of variances was tested using Levene’s test.The statistical significance of differences at baseline was assessed using generalized linear models due to non-normal and left-skewed distributions. Generalized linear models with repeated measures were fitted in order to estimate the dynamics of both continuous and discrete variables throughout the observation period, incorporating a set of explaining variables and being controlled for the patients’ age.p < 0.05 was considered statistically significant. All the statistical procedures were carried out using STATISTICA™, release 14 .A level of p > 0.05. There were no significant differences between the groups at baseline in terms of OSDI score, the Schirmer test, TBUT, the Oxford grade scale, conjunctival hyperaemia, or Meibomian gland parameters. Thus, the studied groups were all clinically well-matched , EGF , fibronectin , VEGF , PDGF AB , NGF , PDGF , and substance P  in PRP compared to AS. The level of TGF-β was undoubtedly higher in AS than in PRP . No significant differences between AS and PRP were observed in the IGF concentration (The concentration of selected GFs in blood-derived eye drops (PRP vs. AS) was analyzed, and the results are presented in ntration .p < 0.0001) in the AS group, whereas the mean value changed from 79 ± 12.71 to 59.81 ± 10.31 at the 3-month visit in the PRP group (p < 0.0001). A reduction in the OSDI questionnaire was noticed in 100% of patients in the PRP group and in 77.3% of the AS group as compared to baseline (p = 0.0031). There was a significant difference in the mean change of the OSDI score between and within the groups . In both groups, patients reported significant improvement in their symptoms after the treatment. The results are shown in The baseline mean OSDI score fell from 79.85 ± 13.12 to 67.07 ± 11.57  after 3 months of therapy.The mean BCVA (logMAR) of all participants improved in the AS group from 0.06 ± 0.07 to 0.04 ± 0.04, (p = 0.0063)  .There was no difference in the baseline mean value of the Schirmer test in the studied groups. In PRP, it was 2.95 ± 2.8 mm, and in AS, 2.91 ± 2.79. However, after 3 months, it improved to 5 ± 2.86 and 4 ± 3.41, respectively. There was a significant difference between the values within the groups after the treatment. Interestingly, there was no significant difference between the two groups after 3 months of therapy .Oral Schrimer test (mm).No significant changes in the mean value of the oral Schirmer test were detected in the researched groups.p < 0.0001, but not between the groups after the treatment, p = 0.1714   .The oral Schirmer test and tear film meniscus did not change significantly.There were no significant changes observed in meibomian gland parameters during and after the treatment between and within the groups .Participants did not report any discomfort or irritation during the treatment period. No temporary or permanent adverse events were noticed during or after the biological product therapy.p = 0.013), TGF-β and PDGF-AB , and PDGF and PDGF-AB . There was a significant positive correlation between TGF-β and EGF , PDGF , PDGF AB , and VEGF  in PRP. In addition, remarkable interactions were found between EGF and FGF , VEGF , PDGF AB , and NGF  in PRP. Moreover, a positive correlation was observed between NGF and EGF , FGF , PDGF, and PDGF-AB  in PRP.In the AS, there were correlations between TGF-β and EGF  in PRP. However, no correlation was noted between PLT and other GFs.There was also a positive correlation between the PLT number and the PDGF level , whereas the statistically significant correlation between the mean OSDI change and the level of VEGF , PDGF , and substance p  was observed only in the PRP group. In the AS group, there was a negative relationship between the OSDI score and the level of TGFβ  and EGF .The relationship between the concentration of the GFs and the mean change of the clinical outcomes was analyzed. There was an interaction between the reduction of OSDI score and the level of NGF in both groups , PDGF , and substance p , but only in the PRP group.Interestingly, the improvement of BCVA was significantly correlated with the concentration of PDGF-AB , EGF , and TGFβ  in the AS group and only with NGF in the PRP group .Regarding the Schirmer test, a correlation was noticed between the levels of FGF , IGF  in the AS group, while in the PRP group it was correlated with IGF  and TGFβ .The mean increase of the TBUT value was correlated with TGFβ  in the AS group and in the PRP group with PGDF , PGDF-AB , and NGF .The mean reduction of Oxford ocular surface staining was connected with the FGF level , PDGF , and PDGF-AB  in the AS group, and with FGF , PDGF , and PDGF-AB  in the PRP group.Regarding the decrease of conjunctival hyperaemia, there was a strong statistical interaction with FGF  and EGF  had an impact on the quality and expressibility of meibomian glands only in the PRP group.Interestingly, the level of substance Haemoderivates appear to be a promising strategy for DE; however, there seems to be no consensus regarding AS and PRP preparation, concentration, or frequency of application. Some authors believe that the higher the platelet count and the higher the number of growth factors inside, the better the treatment results. Thus, the therapeutic benefit of blood-derived products might be explained by their composition, which is very much like natural human tears.Our study is one of the first to compare GF concentrations in 100% AS and 100% PRP and attempt to determine their influence on clinical results after the treatment of DE in pSS patients. So far, no consensus has been achieved regarding the preparation of blood derivatives; thus, products with different compositions and clinical effects are obtained. It was demonstrated that TGF-β concentration was 5 times higher in the AS than in tears; therefore, some authors suggested diluting it to diminish the concentration of these anti-proliferable and pro-fibrotic factors ,23. Howep = 0.0031). Improvement of BCVA was observed in 45.5% of patients in the PRP group and 13.6% in the AS group (p = 0.0063) after 3 months of therapy. The effect on visual acuity could be secondary to the smoothening of ocular surface irregularities or improving subjective symptoms of DE.In this study, a statistically significant increase in BCVA, the Schirmer test, TBUT, meibomian gland parameters, and a reduction in the OSDI scores, Oxford staining, and conjunctival hyperaemia were all observed in each group, but within the groups, the mean changes in BVCA and OSDI scores were more significant in the PRP group. It is worth emphasizing that the patients in the PRP group presented worse BCVA at baseline compared to the AS group, which was not intended and constituted a certain limitation of the study. Thus, the expected improvement in visual acuity may be greater in the PRP group than in the less-affected AS group. That fact was not enough to draw the conclusion that PRP had a statistically significantly better effect on improving visual acuity than AS. The improvement of VA and alleviation of subjective symptoms are the most expected and desired effects of the treatment in this chronic disease, which significantly improves the patient’s quality of life. A reduction in the OSDI questionnaire in this study was noticed in 100% of patients in the PRP group and 77.3% in the AS group compared to baseline  after the treatment was significant in both groups. We do not know exactly how this translates into clinical practice since many factors are responsible for the improvement of the clinical condition and symptoms at the same time, and sometimes a small change in one of them significantly affects the whole condition. Moreover, some relationships between the TBUT improvement and growth factors were also noted, which would be discussed afterwards.However, some of the results achieved by other authors are worth citing. According to the meta-analysis performed by Jongkhajornpong, PRP significantly reduced OSDI score and ocular surface fluorescein staining and increased TBUT more than AS . The finNevertheless, comparing different studies on the subject of PRP and AS was challenging because of the differences in the preparation protocols of the blood products and the different outcome assessments across the studies.The Cochrane meta-analysis of randomized controlled trial reports devoted to the efficacy of 20% AS in comparison to artificial tears suggested that 20% AS eye drops alleviate DE symptoms better than artificial tears, but only for the first period of treatment (2 weeks). The data for the long-term efficacy period was ambiguous . InteresConcerning the application of PRP for the management of dry eye, a successful result with minimal complications has been reported by Alio et al., who observed an improvement of symptoms in 87.5% of participants and decreased corneal fluorescein staining (CFS) in 76.1% of cases. Moreover, 28.8% of patients improved by at least one line of BCVA, and they observed a significant decrease in OSDI scores . Similarp < 0.001) and a significantly higher improvement in TBUT and CFS scores (p < 0.05) after 3 months of treatment with PRP compared to artificial tears. There was no significant difference observed in the post-treatment Schirmer test score between the two groups (p = 0.44) [Additionally, Rawat reported an OSDI score reduction ( = 0.44) . Meibomi = 0.44) . This diSome authors performed studies concerning PRP injection to the lacrimal gland of SSp, obtaining relief from the signs and symptoms of DE ,42.In spite of these encouraging results, there is limited availability of clinical trials proving the superiority of PRP treatment in dry eye disease in SS over autologous serum therapy. Blood derivatives contain bioactive ingredients that are missing in artificial tears. It is believed that PRP provides more advantages than AS eye drops as PRP has a more beneficial composition and its regenerative effect is based on a different concentration of several growth factors. Most of them influence corneal epithelial healing. According to some authors, platelet-derived eye drops provide larger amounts of NGF, PDGF, and fibronectin than AS ,44. PlatHowever, there is still a lack of comparative studies confirming this. According to the Dry Eye WorkShop (DEWS) II report , blood pIn our study, significant differences in the concentration of growth factors between PRP and AS were noted. The level of TGF-β was undoubtedly higher in AS than in PRP. No significant differences between AS and PRP were observed in the concentration of IGF. The observed dependences indicate a complex mechanism of action and synergy in the cooperation of the growth factors with each other. Those data may support the hypothesis that a higher level of growth factors may better relieve the symptoms and enhance corneal epithelial healing. All of this may lead to a reduction in OSDI and ocular surface fluorescein staining. For this reason, PRP has superior efficacy over AS.Our study revealed that there was a higher FGF level in AS correlating with the improvement in the Schirmer test, Oxford staining, and the reduction of conjunctival hyperaemia in both groups, which was in agreement with Xiong research confirming that FGF alleviates DE by modulating the inflammation response and promoting the healing process .Lacrimal glands increase the expression of EGF after corneal epithelium injuries via the trigeminal nerve. Therefore, EGF plays an important role in the healing process by controlling/promoting epithelial cell proliferation and migration and preventing apoptosis. Moreover, EGF increases the number of goblet cells and muc−1 expression, thus contributing to tear film stability  by the mOhashi et al., found a correlation between EGF in tears and the Schirmer test—a decreased level of EGF in tears was observed in SSp. It is widely known that the lacrimal gland’s ability to produce tears is decreased in SS, so the quantity and quality of tears are extremely important to preserve the biological activity of the tear proteins and protect the ocular surface. When the lacrimal gland does not work properly, we can deliver GFs such as EGF to preserve ocular surface function.Despite very advanced dry eye and a higher level of VEGF in AS compared to PRP, we did not observe neovascularization in our patients. This may be due to the inhibitory effect of the other GFs or the balance between the proangiogenetic and antiangiogenetic factors. The concentration of VEGF had a statistically significant correspondence with the reduction in OSDI values in the PRP group and, what is more interesting, with the improvement of the Schirmer test in the AS group. There is still no information on how exactly this GF can influence the abovementioned outcomes. However, some animal studies reported that VEGF was detected in the lacrimal glands of rat models . VEGF caNormally, corneal epithelial cells are permanently renewed by mitosis; corneal neurotrophic factors play a fundamental role in maintaining cornea health. Generally, NGF has a decisive function in the regulation of growth, maintenance, proliferation, differentiation, and survival of corneal nerve and sensory neurons, corneal sensitivity, and even tear production. Moreover, it may be potentially responsible for modulating the proliferation of corneal epithelial cells.It is assumed that the ocular surface and the lacrimal gland are connected by nerves, and their impairment might evoke DE. Deterioration in corneal sensitivity induces a decrease in tear production and suppresses the blinking reflex, resulting in a hyperosmolar environment and inducing inflammation. All of this reduces the expression of epitheliotrophic neuromediators/GFs.p = 0.0016) was noticed. Moreover, a correlation between the level of NGF and the improvement of the Schirmer test value and Oxford staining score in the PRP group was observed. This observation might be explained by the pleiotropic effect of NGF on the ocular surface. NGF is known not only to influence and modulate corneal sensitivity and corneal and conjunctival epithelial proliferation and differentiation, but it also controls immune reactions and stimulates mucin production by goblet cells [p = 0.009) [Cortes et al., measured the NFG level in tears in patients with moderate dry eye syndrome. They observed a negative correlation between NGF and OSDI results . Similaret cells ,53. Likeet cells ,54. This= 0.009) . Nonethep < 0.0001). This growth factor is responsible for cornea health and sensitivity improvement. Therefore, it helps to improve not only local conditions but also subjective symptoms. Nowadays, recombinant human NGF (rhNGF) as an eye drop solution is approved for the treatment of neurotrophic keratopathy. Observational studies results are encouraging; however, due to their high cost, their use is limited [The NGF level was definitely higher in PRP than in AS  [Sang et al., described an increased expression level of substance P in the cornea and conjunctiva that was correlated with the increased CFS scores (< 0.001) . In our PDGF is a modulator of chemotaxis and the cellular mitosis of fibroblasts. Moreover, it stimulates the expression of other factors, especially TGFβ. In PRP, we observed its higher concentration due to platelet degranulation. Interestingly, a negative correlation between the level of PDGF and PDGF-AB and the mean change of BCVA, OSDI score, Oxford staining, and hyperaemia  was founIt is widely known that TGF-β is a multifactorial GF and has antiproliferative properties in a dose-dependent manner. It was demonstrated that TGFβ may suppress corneal wound healing and promote corneal stromal fibrosis—the disorganized extracellular matrix generated by myofibroblasts is responsible for corneal opacities. Therefore, a lower concentration is more desirable in the healing process. A lower level of TGFβ in PRP stimulated corneal regeneration due to weak conversion of stromal fibroblasts to myofibroblasts and scar formation . TGF-β wA statistically significant correlation between many GFs  was observed. This may indicate how complex the ocular surface regenerative process is. The healing process may be based on the interaction and synergistic mechanisms of many biologically active molecules.It is well known that low lipid secretion and a reduction in blinking rate lead to tear film instability and tear evaporation. Ocular surface homeostasis is broken, and corneal epithelium may be injured; thus, meibomian gland function is extremely important. We noticed a negative correlation between meibomian gland parameters and the EGF level and a positive one between substance P, but only in the PRP group. Murtaza et al., reported an improvement of the TBUT and conjunctival hyperaemia in evaporative dry eye disease treated with PRP drops .To understand the biology, molecular mechanism of action, and interaction between GF in PRP and AS therapy, their impact on the ocular surface in DE in SSp should be focused on further studies based on cell cultures or animal models.The literature describes some adverse effects of hemoderivatives, such as eye irritation, discomfort, or even eye discharge. Some studies reported the potential risk of contamination of blood derivatives, but adding sterilizing filters decreased such incidences . AnotherThe storage of blood-derived products at patients’ houses was not controlled and was under the patients’ responsibility. Generally, it was reported that blood products may be used safely in both outpatient and inpatient settings, but under strict preparation and storage protocols ,65. The Lopez-Garcia reported that long-term storage of AS at 20 °C did not change the qualification level of GFs . Anitua Interestingly, Okumura suggested that PRP eye drops stored at 4 °C for 4 weeks had a higher concentration of GFs than AS eye drops .Nevertheless, evaluation of the stability of bioactive molecules should be based on cell cultures or animal models to find out their shelf life. While comparing the clinical outcomes, we found some inconsistencies, in our opinion, due to a huge number of different preparation protocols. Moreover, the quality of blood products may be affected by different preparation protocols, age, gender, race, general disease, diet, and daily routine.Dry eye is an extremely common problem that might also display a chronic character. Over 300 million people worldwide suffer from some degree of dry eye. Blood derivates have a significant positive therapeutic role in the treatment of DE due to the huge quantity of growth factors that promote wound healing at the damaged ocular surface. The treatment with AS and PRP, as presented in our study, resulted in a significant improvement in the signs and symptoms of DE accompanying primary Sjogern’s syndrome.Topical administration of hemoderivates appears to be a very promising strategy for DE treatment. Therefore, understanding their mechanism of action would be highly beneficial.Despite the fact that the blood derivatives differ in composition, they seem to be effective and safe in the treatment of severe DE in pSS patients. The signs and symptoms of DE were reduced in both groups, but only the mean change of OSDI was statistically significant between the groups. A greater reduction in OSDI scores was observed in the PRP group. The obtained results and the composition of haemoderivates may indicate the superiority of PRP in relieving the symptoms of DE in pSS patients compared to AS.


Pre-service teachers should be prepared to face the emotionally demanding situations associated with the profession. The previous literature suggests that two variables are important for managing teaching demands: emotional intelligence (EI) or the ability to perceive, facilitate, understand, and manage emotions and implicit theories (ITs). ITs refer to the beliefs about the malleability of various life domains. Individuals can be divided into incremental theorists  and entity theorists (attributes are fixed).This study aimed to evaluate the influence of ITs of intelligence and EI on self-report and ability EI in a sample of female preschool pre-service teachers.M = 21.27, SD = 4.72) were assessed on ability EI (performance and self-report instruments), ITs of intelligence, ITs of EI, age, and parental education.In total, 224 participants (In our sample, incremental EI—but not intelligence—theories predicted higher scores on self-report and ability EI. In particular, being an incremental theorist of EI predicted 11 and 20% of the variance of the global EI and the managing branch of the ability EI, respectively.Our findings suggest the importance of ITs of EI for pre-service teachers’ emotional intelligence and open the door to implementing ITs of EI training in this population. These theoretical and practical implications are discussed. Differeperformance-based ability model of EI. Self-reports evaluate individuals’ subjective perception of their EI and are part of the so-called self-report ability model. Another way to operationalize EI is by using the self-report mixed models, which consider EI as a combination of social and emotional skills  have found incremental theorists to show better academic and professional achievement, intrinsic motivation, stronger learning goals, higher tendencies toward making an effort, and better mental health and wellbeing . In addiIndividuals also differ in the way they perceive the malleability of EI, that is, in their IT of EI (ITEI). ITEI has been scarcely studied, but a related starting point could be to look at IT about emotions. Being an entity theorist of emotions has been linked to lower social support, less adaptative emotional regulation abilities, and reduced wellbeing; the latter being demonstrated not only in the general population but also in pre-service teachers .Regarding ITEI, Preschool is a benchmark for young learners as it implies the beginning of the “school age.” This is an important period in which teachers’ EI acts as an early model for emotional management . TherefoH1. A weak relationship will be found between the self-report ability model and the performance-based ability model of EI.H2. ITEI will have a stronger influence than ITI on both models of EI, after controlling for sociodemographic variables.H3. Incremental theorists of EI will have higher EI scores according to the performance-based ability model, after controlling for sociodemographic variables.H4. Incremental theorists of EI will have higher EI scores according to the self-report ability model, after controlling for sociodemographic variables.2.2.1.M = 21.27, SD = 4.72). All the participants were Spanish speakers. They took part in the study in exchange for course credits. All participants provided written informed consent after being told of the details of the study. The study was carried out in accordance with the Declaration of Helsinki  with a split-half reliability of 0.93, and the test–retest reliability of the total MSCEIT of 0.86 after 3 weeks .Trait Meta-Mood scale-24 TMMS-24; . The sel2.2.5.Participants were asked about their father’s and mother’s level of education through a self-report question with three response categories: primary studies, high school, and college education.2.3.Participants completed the questionnaires through an online platform (LimeSurvey) provided by the university, where blank responses were not allowed in order to avoid missing data.2.4.Descriptive statistics, Pearson’s correlations, and regression analyses were carried out using SPSS version 24.0 . First, descriptive analyses and Pearson’s correlations were computed for the study variables. Second, stepwise multiple regressions were run to test whether implicit theories of intelligence and EI were a significant predictor of the perceived and ability EI of female pre-service teachers while controlling for age and the parental education status. Five regression models were run to test the association between implicit theories of intelligence and EI and MSCEIT: total (Model 1), perceiving (Model 2), facilitating (Model 3), understanding (Model 4), and managing (Model 5). Additionally, three regression models were run to test the association between implicit theories of intelligence and EI and TMMS: attention (Model 6), clarity (Model 7), and repair (Model 8). The control variables of age and parental education status were included as predictors in each model.3.r = 0.17, p < 0.01) and MSCEIT managing , and ITEI and MSCEIT total , MSCEIT managing , MSCEIT understanding , and MSCEIT facilitating . Furthermore, concerning TMMS, we found a positive relationship between ITI and attention , clarity  and repair  and between ITEI and attention  and clarity . In addition, ITI was positively associated with the ITEI . Finally, regarding the relationship between MSCEIT and TMMS, we found only a positive relationship between MSCEIT total and clarity  and between MSCEIT managing and clarity .Descriptive statistics and Pearson’s correlations of the study variables are displayed in R2 = 0.02) and MSCEIT understanding (R2 = 0.04) but was the greatest for MSCEIT total (R2 = 0.11) and particularly MSCEIT managing (R2 = 0.20). In contrast, the best predictor for MSCEIT perceiving (Model 2) was parental education status, with a small unique contribution (R2 = 0.04).The results of five linear stepwise regressions to test the association between ITI, ITEI, and MSCEIT total and the four branches are reported in R2 = 0.01) and clarity (R2 = 0.02). In contrast, the best predictor for repair (Model 8) was ITI with a small contribution (R2 = 0.02).The results of three linear stepwise regressions testing the association between ITI, ITEI, and TMMS dimensions are reported in 4.The present study aimed to analyze the influence of ITI and ITEI on EI. Specifically, we wanted to determine to what extent incremental or entity theories of intelligence and EI impact the EI ability, assessed through both objective performance and subjective self-report measures.Starting with our target variables, and as expected (H1: A weak relationship would be found between the self-report ability model and the performance-based ability model of EI), we only found weak relationships between some of the scales of the performances and the self-report ability models of EI. Specifically, we only found significant positive correlations between the clarity subscale of the TMMS and the total and the understanding branch of the MSCEIT. This is congruous with previous literature and emphasizes that although both models are conceptualized in the same way, they do not cover the same construct . TherefoConcerning the main aim of our study, that is, analyzing the influence of ITI and ITEI on EI scores (via both self-report and performance measures), we hypothesized (H2) that ITEI would have a stronger impact than ITI on these variables. Correlational analyses revealed that ITI was positively and weakly related to both the total and managing branches of the performance-based ability model of EI and the three subscales of the EI self-report ability model. ITEI correlated positively and weakly with the total and the facilitating and understanding branches of the MSCEIT and moderately with the managing branch, that is, with the performance-based ability model. In addition, weak positive correlations were found between ITEI and the attention and clarity subscales of the TMMS or the self-report ability model. To determine the variable that more strongly influences EI, eight multiple regression analyses were carried out. These analyses showed that ITEI predicted the total and the managing, facilitating, and understanding branches of the MSCEIT and the attention and clarity subscales of the TMMS beyond the control variables and, more importantly, beyond ITI. Only the repair subscale of the TMMS was better predicted by the ITI. This indicates that, as hypothesized, being an incremental theorist of EI influences the EI of the pre-service teachers to a greater extent than believing intelligence is malleable. Overall, this result is consistent with those of previous studies with adolescents and the general population ,b. TheseThe present findings also support our H3, that is, incremental theorists of EI showed higher scores on the performance-based ability model of EI. This was predicted for all the EI branches except the perceiving branch of the MSCEIT. Nonetheless, this result was particularly significant for both the total and managing branches of EI, for which ITEI explained 11 and 20% of the variance, respectively. This result is consistent with the findings of Therefore, taken together, the present results suggest that ITEI had a greater influence on the performance EI scores than the self-perceptions of the pre-service teachers, and principally, this influence is on the ability to regulate emotions. It is particularly interesting to note that the managing branch was most influenced by ITEI, given that the ability to regulate emotions is of central importance for creating adaptive responses in emotional situations . FurtherThe lower influence of ITEI on self-perceived EI compared with performance measures is inconsistent with previous studies where the opposite pattern of results has been reported ,b. HowevThis study has some limitations. First, the correlational nature of the study prevents us from making any causal inferences. Thus, future investigations should focus on causally analyzing the impact of ITEI on both the performance and self-report EI scores by, for instance, testing the efficacy of ITEI programs. Second, our sample comprised only female adults, and thus, the results cannot be generalized to male adults. The recruitment of our sample reflected the higher proportion of female students who were enrolled in the teacher training courses. Finally, only 17 male adults were included in our sample, and we decided to remove them from the analysis since they only accounted for 7% of the sample, leading to underpowered analyses . However5.In conclusion, in a sample of female pre-service teachers, ITEI appears to be associated with managing emotions (which is the most complex branch of the EI model) while showing a weaker relationship with self-perceived EI. These results have important theoretical and clinical implications. Specifically, our results contribute to the literature on ITs by suggesting the importance of ITEI for the EI of pre-service teachers. This also opens the door to new research studies in teachers and pre-service teachers of other grades . Finally, our results are of clinical relevance since they could inform the development and implementation of ITEI training programs in order to evaluate their impact on the EI of female pre-service teachers. Such training could be included in the career curriculum.https://riuma.uma.es/xmlui/handle/10630/22905.The original contributions presented in the study are publicly available. This data can be found here: The studies involving humans were approved by the Research Ethics Committee of the University of Malaga. The studies were conducted in accordance with the local legislation and institutional requirements. The participants provided their written informed consent to participate in this study.MG-C: Conceptualization, Data curation, Funding acquisition, Investigation, Methodology, Resources, Validation, Writing – original draft. RC: Conceptualization, Formal analysis, Funding acquisition, Investigation, Writing – review & editing. PF-B: Conceptualization, Formal analysis, Funding acquisition, Investigation, Methodology, Project administration, Supervision, Writing – review & editing.


Necrotizing soft tissue infection (NSTI) of the breast is an extremely rare event in surgical practice. It is considered the most aggressive form of soft tissue infection and a true surgical emergency. It is also associated with a high risk of mortality if not diagnosed promptly. Few cases have been documented in the literature; the exact etiology and risk factors vary from those involving the limbs, trunk, and perineum. Early recognition, prompt surgical treatment, and broad-spectrum antibiotic therapy are crucial for reducing morbidity and mortality. These reports present cases of NSTIs in breasts with unique etiologies and challenges in their management. Necrotizing fasciitis of the breast is a clinical entity infrequently described in surgical texts and rarely encountered in practice. Unlike other necrotizing soft tissue infections (NSTIs), this entity occurs due to a variety of causes, including local trauma, breast infarction, and topical applications. Mortality rates have been reported as high as 73% but can be reduced with early diagnosis and the rapid initiation of appropriate multidisciplinary strategies .We present four cases of breast NSTI, each with unique etiologies and challenges along with their management pathway. We seek to discuss the incidence of this rare pathology, its associated microbiology, and the options for surgical management.A 60-year-old female presented to the Accident and Emergency Department with a “ruptured right breast.” She was diagnosed approximately ten months prior with triple negative, locally advanced (T3N2M0) breast carcinoma, which progressed to T4bN2M0 disease while on neoadjuvant chemotherapy . She wasOn assessment, the patient was hypotensive (blood pressure 94/69 mmHg), tachycardic with a pulse of 140 beats per minute, respiratory rate of 34 breaths per minute, and oxygen saturation of 98% in room air. There was a large ulceration of the tumor at the lateral aspect of the right breast with a foul odor and significant drainage of turbid fluid . The rig9/L), hyponatremia (129 mmol/L), hypokalemia (3.3 mmol/L), acute kidney injury, and serum creatinine 2.2 mg/dL. Non-contrast computerized tomography (CT) showed a significant amount of gas throughout the right breast, with an invasion of the tumor into the pectoral muscles  5.5 g/dL], leukocytosis .At the examination, the patient appeared unwell, with a blood pressure of 133/56 mmHg, a pulse of 133 beats per minute, a respiratory rate of 30 breaths per minute, and an oxygen saturation of 99% in room air. There was extensive erythema in the medial aspect of the left breast, involving the nipple–areola complex and skin necrosis at the site of the sebaceous cyst . There w9/L) and acute kidney injury (serum creatinine 1.6 mg/dL). A chest X-ray appeared suspicious for subcutaneous emphysema of the left chest wall  presented to the Accident and Emergency Department with one week of history of pain, swelling, redness, and discharge of pus from her left breast. There was no history of trauma or any insect bites on her breast, and she complained of a mild fever for 48 hours. She completed her second cycle of chemotherapy and was on rivaroxaban for her bilateral PE.On examination, the patient appeared unwell, with a blood pressure of 142/77 mmHg, a pulse of 128 beats per minute, a respiratory rate of 26 breaths per minute, and an oxygen saturation of 95% in room air. There was mild erythema of the lower inner quadrant of the left breast with a 10 cm × 10 cm tender mass, no palpable crepitus but purulent discharge , and the9/L), Hb of 7.2 gm/dL, and normal platelet and kidney function tests. A CT scan of the chest confirmed extensive gas within the left breast , Hb of 10 g/dL, a normal platelet count, and a kidney function test.Her laboratory investigation revealed marked leukocytosis . These infections typically involve the perineum, lower limbs, and torso, involving extensive soft tissue destruction along with systemic signs of sepsis. They are often associated with significant mortality and morbidity, particularly because of diagnostic and treatment delays , 3.n = 2) and diabetes (n = 1), and one had no comorbid condition except obesity.Originally described in the surgical literature by Shah et al., necrotizing fasciitis of the breast is a rare phenomenon. This entity is often misdiagnosed as an abscess, cellulitis, or inflammatory breast cancer, reflecting difficulties in recognition and leading to treatment delays . PredispBreast gangrene, in particular, has been reported in trauma, thrombophlebitis, pregnancy and lactation, anticoagulation, and topical belladonna application , 5. HoweCommon manifestations of NSTIs of the limbs, trunk, and perineum include skin erythema, edema, and pain out of proportion to exam findings in up to 75% of cases. Additionally, up to 50% of patients have palpable crepitus with skin bullae or necrosis , like ouNSTIs of the breast, however, can prove to be a diagnostic challenge for surgeons. An extensive blood supply to the breast contributes to a delay in skin manifestations, with most patients (including the cases presented here) presenting days to weeks after the onset of symptoms . FurtherClostridium, Klebsiella, or Vibrio species. Type IV infections, on the other hand, are fungal in nature [E. coli only ; the second patient's culture placed her in the type I category, whereas the third patient grew both E. coli and Klebsiella.NSTIs are broadly categorized into four types. Type I involves mixed aerobic and anaerobic bacterial infections, often presenting as fulminant sepsis. Type II infections are generally mono-microbial  and often occur in patients with significant risk factors , 3. Typen nature . A 2020 n nature . InteresIn addition to clinical appearance, the presence of systemic signs of toxicity lends itself to the diagnosis of NSTIs. The Laboratory Risk Indicator for Necrotizing Fasciitis (LRINEC score) is a clinical tool that uses laboratory values and clinical signs as adjuncts in the diagnosis. Here, a score of 6 or more helps differentiate necrotizing from non-NSTIs . The LRIVarious imaging modalities have been used to aid in the diagnosis of NSTIs, especially in cases where the diagnosis may not be immediately apparent. Ultrasound can detect the thickening of the deep fascia and fluid collections. CT can identify fat stranding and subcutaneous gas characteristics in these infections. The CT scan for our first and second patients revealed the presence of gas throughout the breast parenchyma, which supported the need for a mastectomy due to the extent of gas; however, in our third patient, the gas was limited to the lower inner quadrant of the breast, which supported a wide debridement instead of a mastectomy. The additional benefit of CT in our first case was that it provided some degree of the staging of her breast carcinoma and enabled a decision for radical versus modified radical mastectomy. This finding also supported the aggressive nature of her disease: progression to pectoral involvement despite multiple lines of neoadjuvant chemotherapy.On the other hand, the question of whether a CT is necessary was raised in the management of our second case. The presence of characteristic clinical and systemic signs of NSTI, along with suspicious X-ray features, suggested that a CT was not necessary for diagnosis. No diagnostic imaging was performed when it was first described by Shah et al., likely because of the limited availability of CT at that time . HoweverOther adjuncts, like Magnetic Resonance Imaging, have been described, especially as they provide a more detailed view of the extent of soft tissue infection. However, its specificity is low (46–86%) and, in a resource-limited setting like ours, is not readily available at times of surgical emergency .A multidisciplinary approach to treating NSTIs of the breast is strongly recommended and involves input from breast surgeons, plastic surgeons, critical care specialists, microbiologists, and radiologists. This provides the platform for better patient outcomes with clear planning and decisive perioperative management . Shah etMarks et al. subsequently suggested an abbreviated diagnostic and management triad based on a modification of this 6-point system .Early debridement and antibiotic coverage significantly reduce the morbidity and mortality associated with NSTIs. In cases involving the breast, though, reports supported mastectomy as the most commonly performed procedure (52% of cases in two systematic reviews) .n = 1) due to the rupture of neglected giant breast cancer, and the mortality rate was 50%; however, this was not related to the necrotizing fasciitis of the breasts. One of our patients (the first case) was diagnosed due to the advanced stage of her breast cancer (from brain metastasis) after 3–4 months of surgery, and in the third case, it was due to her pre-existing bilateral PEs after 4 weeks of surgery.The mastectomy rate in our series was 25% , local flap advancement (as in our second case), local breast tissue rearrangement (an option for our third and fourth cases) with or without contralateral symmetrization (as in our fourth case), and pedicle flap reconstruction .A prompt diagnosis of breast NSTIs is crucial for saving a life as well as potentially salvaging the breast. A diagnosis can be readily made once clear clinical signs are apparent, but when not, radiologic adjuncts can be beneficial in achieving a diagnosis and guiding surgical treatment. These patients require aggressive resuscitation and control of infection with debridement until healthy, viable tissue is encountered. This often requires a mastectomy. Re-exploration and re-debridement are necessary until all infection is cleared, and reconstruction may then be considered once it is feasible.


Pancreatic β-cells and α-cells have been found in the murine extrahepatic biliary ducts but not in the gallbladder. However, there has been no information reported in the specialized literature about the presence of glucagon- and insulin-expressing endocrine cells in porcine bile ducts and gallbladder.We aimed to perform an immunohistochemical study to identify glucagon- and insulin-positive cells and their distribution in the porcine extrahepatic biliary ducts and gallbladder.ductus hepaticus communis), common bile duct (ductus choledochus), cystic duct (ductus cysticus), and gallbladder (vesica fellea) of male pigs. Chromogranin A was used as a typical marker for endocrine cells.The immunohistochemical method was used to detect the presence and distribution of glucagon- and insulin-positive endocrine cells in the common hepatic duct (The density of chromogranin A-, glucagon- and insulin-positive cells per field was the largest in the common bile duct, followed by the common hepatic duct, cystic duct, and gallbladder. The three types of endocrine cells showed specific localization in the superficial and deep glands of the studied organs.The distribution of glucagon- and insulin-immunopositive endocrine cells in the porcine extrahepatic biliary tract was established for the first time as a new source of these hormones. The presence of α- and β-cells in the epithelium of extrahepatic bile ducts can be applied in treatment of diabetes, taking into account the possibility to reprogram the biliary epithelium to mentioned pancreatic endocrine cell types. Due to the similarity in body weight, anatomy, and physiology between pigs and humans, the pig has become a model for morphological, physiological, biochemical, and genetic investigations .The anatomical and physiological similarities of organs such as the liver, pancreas, kidney, and heart have also made the pig a major species of interest as an organ donor for xenotransplantation procedures , 3.Despite minor morphological differences, swine are the ideal species for liver xenotransplantation. Modern medical advances have enabled xenotransplantation of the liver from genetically modified pig donors. New medical techniques could make xenotransplantation an important method to solve the problem of providing livers from human donors .ductus hepaticus communis, DHC), the common bile duct , and the cystic duct . The gallbladder  collects bile and releases it through the cystic duct into the common bile duct  and insulin (Ins+Cs)-immunopositive endocrine cells in the wall of porcine DHC, DCH, DC, and VB for better understanding of the pathophysiology and treatment of diabetes.We aimed to describe for the first time the localization and density of glucagon-  (92–100 kg) supplied from a single commercial farm, subjected to a standard age-appropriate diet and slaughtered in a regulated abattoir approved by the Bulgarian Food Safety Agency, and was funded by the Scientific Project number 13/2017, Medical Faculty, Trakia University, Stara Zagora, Bulgaria. Six porcine livers, together with the gallbladder, cystic duct, common hepatic duct, common bile duct, and duodenum, were collected from pigs intended for meat consumption at the slaughterhouse. Tissue samples were obtained from the common hepatic duct (DHC), the initial part of cystic duct (DC) near the gallbladder’s neck, the initial part of the common bile duct (DCHO) near the junction with the cystic duct, the intramural part of common bile duct (DCHI), and the gallbladder’s neck immediately after slaughtering and fixed in a 10% aqueous solution of formalin.Routine histological techniques were used to process the material and to obtain serial paraffin sections that were stained with hematoxylin and eosin to exclude the presence of pathological findings. Another part of the sections was processed immunohistochemically for detection of glucagon and insulin expression.In this work, the ABC (avidin-biotin peroxidase complex) technique was performed. Serial tissue sections with 5 μm thickness were washed in 0.1 M PBS and placed in 1.2% hydrogen peroxide in methanol for 30 min. Antigen retrieval in buffer (pH 9.0) was done for 20 min. Between steps, sections were washed with an EnVision Flex Wash Buffer, then incubated in a humidified chamber overnight at 4°C with primary antibodies: glucagon mouse monoclonal antibody  SC-514592, Santa Cruz Biotechnology, Dallas, TX, United States), insulin mouse monoclonal antibody , SC-8033, Santa Cruz), and chromogranin A rabbit antibody (PA 0430) (Leica Microsystems Inc.), which were ready to use. The immune reaction was visualized with diaminobenzidine. Three serial sections on a slide were stained consequently with glucagon-, insulin-, and chromogranin A antibodies. Three slides per animal were used.PBS is used instead of primary antibody as a negative control.p-values of less than 0.05 were considered statistically significant. The data are presented as mean ± SD.The number of endocrine cells was estimated on three microscopic fields X100 from three sections of the DHC and extra- and intramural parts of the DCH, DC, and gallbladder’s neck for each antibody and per each animal. The data for endocrine cell density (number of endocrine cells per field and per cross section of a gland) were processed by Graph Pad Prism 6 for Windows  via one-way ANOVA followed by the Tukey–Kramer post-hoc test. +Cs) in the glands of the gallbladder and extrahepatic bile ducts  was used as a marker for endocrine cells to detect all endocrine cells in the studied organs. In this manner, we identified the largest number of endocrine cells  was significantly larger than Glu+Cs . In the DCHI, the number of Ins+Cs  and Glu+Cs  was similar. In DC, the density of Ins+Cs was 89% from all ChrA+Cs but of Glu+Cs – 60%. In VB, the number of Ins+Cs, Glu+Cs, and ChrA+Cs was the same.The density of Ins ChrA+Cs . In the +Cs and Glu+Cs per field was detected in the DCHI, followed by the DCHO, DC, DHC, and VB  in the VB were of closed type. However, in the extrahepatic bile ducts, both open and closed type of ChrA+Cs were observed. Ins+Cs and Glu+Cs were represented by closed type only in the DHC, DCHO, DC, and VB. In the DCHI, both open and closed Ins+Cs and Glu+Cs types were present and their number was equal. Therefore, the secretory products of open cell type acted locally or on distant target cells through the bloodstream. The cells of closed type were localized between other epithelial cells which means that, like intestinal closed cell types, they do not interact directly with luminal substances (In the present study, elongated open-type endocrine cells and oval closed-type endocrine cells were observed, which correlated with the findings of a number of authors regarding the shape of intestinal endocrine cells . It is kbstances .The presence of glucagon- and insulin-positive endocrine cells in porcine bile ducts and gallbladder can be explained by studies by other authors , 39 that+ liver cells are represented by small bile ducts, hepatoblast-like progenitors in the periportal area, or peribiliary glands within larger bile ducts. Thus, Banga et al. (in vivo reprogramming of bile duct cells to a β-cell-like phenotype that can relieve diabetes in adult individuals.According to Banga et al. , Sox9+ la et al.  providedLund et al.  revealedWe supposed that the presence of glucagon-producing cells in porcine extrahepatic bile ducts and gallbladder was probably related to the ability of this hormone to perform the same function in these organs like gut. Based on the results of our research, we hypothesize that there is a functional connection between the extrahepatic bile ducts and the islets of Langerhans which can be defined as a biliary-islet axis, similar to the functional connection between the intestine and the islets of Langerhans named by Fehmann et al.  the enteThe original distribution of our study was identification of porcine β- and α-cells in porcine extrahepatic bile ducts and gallbladder. The largest number of glucagon- and insulin-positive cells was detected in the intramural part of the DCH, followed by its extramural part, the DC, DHC, and VB. These findings defined biliary endocrine cells as components of the gastroenteropancreatic endocrine system and as another source of glucagon and insulin, which can be used in the development of new approaches in the treatment of diabetes mellitus.The raw data supporting the conclusions of this article will be made available by the authors, without undue reservation.The animal study was approved by Ethics committee of the Bulgarian Food Safety Agency. The study was conducted in accordance with the local legislation and institutional requirements.IS: Conceptualization. IS, MG: Methodology. IS: Software, Formal analysis. IS: Investigation. IS: Data curation. IS: Writing - original draft preparation. IS, SS, and MG: Writing - review and editing. IS and SS: Visualization. IS: Supervision. IS: Project administration. All authors have read and agreed with the published version of the manuscript.


We aimed to evaluate the effects of integrated Chinese and Western medical therapeutic modalities on clinical prognosis in a population with stable angina pectoris (SAP) of coronary heart disease (CHD).In a prospective cohort study, 732 patients with SAP of CHD hospitalized in the Integrated Cardiology Unit of the China-Japan Friendship Hospital From October 2020 to October 2021 were included. The patients were divided into integrated treatment and conventional treatment groups according to whether they had been taking Chinese medicine for more than 6 months per year. The occurrence of composite cardiovascular events (CVEs), including cardiac death, non-fatal myocardial infarction, revascularization, stroke, all-cause death, and readmission due to angina attack, heart failure, or malignant arrhythmia, was recorded during follow-up. The effects of different treatment modalities on prognosis were evaluated using univariate and multifactorial logistic regression. Logistic regression models were evaluated using receiver operating characteristic (ROC) curves. In sensitivity analysis, the correlation between treatment modality and outcome events was corrected by rematching the two groups of patients using the propensity score matching (PSM) method.P = 0.037). Covariate correction by multimodal multifactorial logistic regression revealed a lower risk of CVEs in patients receiving integrated therapy compared with conventional therapy . Moreover, a history of renal insufficiency  and a higher Gensini score  were risk factors for the development of CVEs. Model evaluation showed that C-statistic = 0.955 and area under the ROC curve (AUC) = 0.955. After PSM correction, the results still showed that integrated Chinese and Western medical treatment reduced the occurrence of CVEs in patients compared with Western treatment alone .The data from 690 patients were included in the analysis, with 327 patients in the integrated treatment group and 363 patients in the conventional treatment group. CVEs occurred in 19 patients (5.8%) in the integrated treatment group and 37 patients (10.2%) in the conventional treatment group. The proportion of outcome events was significantly lower in the combination treatment group than in the conventional treatment group (Integrated treatment based on Chinese and Western medicine might improve the prognosis and reduce the risk of CVEs in this disease population.http://www.chictr.org.cn/showproj.aspx?proj = 30170.China Clinical Trials Registry, ChiCTR1800017891, Registered 20 August 2018,  SAP of The aim of this study was to further investigate the possibilities to improve the prognosis of the population with SAP of CHD. We adopted a prospective cohort study to assess the incidence of CVEs in real-world patients with SAP of CHD, treated with integrated Chinese and Western medicine or conventional Western medicine, and examine whether there are differences in the risk of CVEs.2.2.1.This study is a prospective cohort study and was conducted following the guidelines laid down in the TRIPOD statement . A totalThis study was approved by the Clinical Research Ethics Committee of the China-Japan Friendship Hospital (2020-114-K73), conducted in accordance with the Declaration of Helsinki, and registered with the China Clinical Trials Registry (ChiCTR1800017891). All patients signed informed consent and gave permission to use the data for further clinical studies.2.2.Patients were diagnosed in reference to the “1979 Nomenclature and Diagnostic Criteria for Ischemic Heart Disease”  jointly 2.3.2.3.1.General information was collected, including demographics , other relevant medical history , and personal habits . Physical activity was reported by the patients. High physical activity was defined as moderate-intensity physical activity for more than 60 min per day.The 19-item Seattle Angina Questionnaire (SAQ) , low-density lipoprotein–cholesterol (LDL-C), lipoprotein a (LP-a), glycosylated hemoglobin (HbA1c), blood urea nitrogen, and serum creatinine (Scr) were tested at the Laboratory Department of the China-Japan Friendship Hospital. The last coronary angiography results were retrieved from the Medical Record System of the China-Japan Friendship Hospital to record the lesions of the vessels. The Gensini scores were calculated according to the Guidelines for Calculating Gensini Scores . All pat2.3.3.β-blockers, calcium antagonists (CCB), and lipid-lowering drugs  was recorded.No medication interventions were made in this study, and the treatment regimens were based on the prescriptions issued by the patients' primary care physicians. Usage of antiplatelet agents , anticoagulants , antimyocardial ischemic drugs , vasodilators (nitrates), ACEI/ARB On the basis of the above, some patients received traditional Chinese medicine treatment according to their own will or their doctor's recommendation. Traditional Chinese medicine included Chinese herbal soup and proprietary Chinese medicine, which were individually prescribed by the attending physician. Patients who continued to take Chinese medicine for more than 6 months per year during follow-up were recorded as “combined therapy”; otherwise, they were recorded as “conventional therapy”. All patients maintained continuous and regular treatment, with specific medications adjusted at follow-up visits.2.3.4.Follow-up was performed by an independent investigator, with outpatient or telephone follow-up visits every 6 months after patients were enrolled in the study to collect information over time until the end of the 12-month study period. Patients whose medication regimen changed significantly during the follow-up period (three or more changes in the Western medication regimen), patients who withdrew on their own, patients who could not be contacted, and patients who were unable to cooperate in completing the follow-up were excluded from further analysis.CVEs were used as the primary endpoint events in this study, including cardiac death, non-fatal myocardial infarction, revascularization, stroke, all-cause death, and readmission due to angina attack, heart failure, or malignant arrhythmia.2.3.5.AR events were defined as any adverse medical events that occurred during the follow-up period, whether they were causally related to drug use. During the follow-up, a small number of patients presented difficulties in stool, fatigue, liver function impairment, gastrointestinal bleeding, skin purpura and other unexpected events, which were uniformly recorded as AR.2.3.6.A total of 35 variables were included in the research as factors that could potentially influence prognosis. Ultimately, 723 patients with SAP of CHD were included, in accordance with the principle of approximately 10 patients per group for each variable in the regression analysis .2.4.t-test was used for normally distributed data, the rank sum test  was used for non-normally distributed data, and the chi-square test was used for count data.Statistical analysis and visualization were performed in R (V4.1.1). Normally distributed data are expressed as mean ± standard deviation, non-normally distributed data are expressed as median (interquartile range), and count data are expressed as percentages. When comparing the means of two sample groups, the independent samples P < 0.05 was considered statistically significant.Patients who completed the follow-up were selected for per-protocol analysis (PPS). Multiple interpolation was performed using the MICE package in R for variables with missing values <20%. Logistic regression analysis was used to observe whether different treatment modalities had an impact on the outcome events. Univariate and multifactorial logistic regression was used to evaluate whether different treatment modalities affected patient prognosis. Logistic regression models were evaluated based on consistency indices (C-statistic) and time-dependent receiver operating characteristic (ROC) curves. At last, sensitivity analyses were performed. The two groups  were rematched based on propensity score matching (PSM) to minimize bias in the observed values and to correct for confounding factors, followed by a corrective test for the association between treatment modality and outcome events. Considering the variables of age, sex, BMI, physical activity, smoking, alcohol consumption and others, the score was calculated using a logistic regression model. A 1:1 ratio matching between the two groups was performed to maximize the PSM based on the nearest neighbor method. 3.3.1.We excluded 33 (4.6%) patients with incomplete follow-up data. Follow-up statistical analyses were performed in the final PPS with 690 patients for whom complete data were available.P < 0.001), a lower percentage of hyperlipidemia (P = 0.034) and lipid-lowering medication (P = 0.001), and lower Scr values (P = 0.005) compared with the conventional treatment group. This reflects the clinical reality of the treatment of patients with SAP of CHD, where clinicians tend to use surgical methods (percutaneous coronary intervention) first to solve the main problem in patients with more complex conditions and start the combined intervention with herbs only after clarifying the overall situation, to avoid possible complications and adverse effects. The incidence of AR during treatment was also lower in the integrated treatment group than in the conventional treatment group (P = 0.014).The number of patients was 327 in the integrated treatment group and 363 in the conventional treatment group. Baseline characteristics are shown in 3.2.P = 0.037).At the end of the 12-month follow-up period, 56 (8.1%) CVEs were reported, with 19 (5.8%) CVEs in the combination treatment group and 37 (10.2%) CVEs in the conventional treatment group. The proportion of outcome events was significantly lower in the combination treatment group than in the conventional treatment group (β-blockers) and AR; Model III was further corrected for the continuous variables of examination results  and SAQ scores. Model III was used as the main study model. The results of the logistic regression tests are shown in We used univariate logistic regression to analyze the relevance of each variable to the outcome event . FurtherP < 0.001) and that combination therapy reduced the risk of CVEs compared with conventional therapy . In addition, physical activity  and regular use of antiplatelet medication  reduced the risk of CVEs, whereas a history of renal insufficiency  and a higher Gensini score  were risk factors for the development of CVEs. Higher Exertional capacity scores  and Treatment satisfaction scores  on SAQ were associated with a lower risk of CVEs.After multivariate correction in Model III, we found that treatment modality was an independent factor of CVEs in patients with SAP with CHD (P = 0.023). Meanwhile, there was no significant change in the incidence of CVEs in patients with or without AR .It is the goal of TCM to increase efficacy and reduce side effects, so we paid extra attention to the occurrence of AR during the follow-up period, and built a regression model in the similar way to determine whether different treatment methods are related to AR. The results showed that combined therapy could reduce the risk of AR  had the highest proportion of CVEs, followed by all-cause death . Therefore, we analyzed the outcomes separately, as a secondary focus. We found no significant difference in the type of outcome between treatments . Different treatment methods, renal insufficiency, antiplatelet, Gensini score, Exertional capacity scores and Treatment satisfaction scores are the influencing factors , indicating that the overall result of the model was statistically significant. The discrimination of the model was tested and we found that C-statistic = 0.955. We used ROC curves based on the proportion of CVEs during follow-up (The logistic regression model (Model III) was statistically evaluated with a negative 2-fold log-likelihood value, and the Omnibus test showed that ollow-up  for mode3.4.Real-world data may suffer from baseline bias. We observed differences in alcohol consumption, hyperlipemia, usage of lipid-lowering drugs, and Scr between the two groups . We perfP = 0.108). We considered that the differences between different groups were masked by confounding factors in the single-factor Chi-square test. The results of the sensitivity analysis (regression methods) were similar to those produced by the main model (Model III), showing a statistically significant impact of treatment modality on the occurrence of CVEs, with the combination of Chinese and Western medicine acting as a protective factor against CVEs .Analysis process of the data in 4.The effectiveness of TCM in the treatment of cardiovascular disease has been widely demonstrated in clinical practice –16. DespClinical characteristics showed that elderly men formed the majority of patients included in this study, most of whom had a combination of hypertension, diabetes mellitus, carotid atherosclerosis, and hyperlipidemia, with a few sufferings from stroke and renal insufficiency, suggesting the complexity of the general profile of the population in patients with SAP of CHD. Moreover, we observed that most patients' lifestyles could be improved, in terms of insufficient daily physical activity, consumption of tobacco and alcohol, etc. In particular, smoking, alcohol consumption, and excessive obesity are serious risk factors for cardiovascular disease, accelerating disease progression and increasing the risk of death –19. ThisP < 0.001), a lower proportion of hyperlipidemia (P = 0.034) and lipid-lowering medication use (P = 0.001), and lower Scr values (P = 0.005) were observed in the integrated treatment group than in the conventional treatment group. It was considered that the uneven distribution of covariates between the two groups occurred due to the non-randomized design used in this study. However, the results also reflect the current treatment principles of integrated Chinese and Western medicine therapies. For patients with more complicated conditions, to reduce adverse effects of integrated medications, clinicians give priority to standardized treatment protocols to solve the main problem and give Chinese herbs after clarifying the overall condition of the patient.In the baseline information of the two groups, a higher proportion of alcohol consumption (P < 0.001). We also revealed that physical activity, regular use of antiplatelet drugs, history of renal insufficiency, a higher Gensini score, and some SAQ items were independently related to disease prognosis. Available studies (In this study, composite CVEs were selected as an outcome indicator , and we  studies , 22 have studies . Renal f studies , and the studies –27 and “ studies , and pat studies . SAQ sco studies , 31. OurIn clinical trials, the interventions, such as innovative operations and complex combined interventions, are often specific and cannot be randomized. PSM  is commoOur real-world prospective cohort study adequately preserved the individualized treatment characteristics in an objective and realistic clinical treatment environment. The relatively broad inclusion and exclusion criteria may increase the heterogeneity of the included population. In addition, the group of combined therapy is integrated with individualized Chinese medicine treatment on the basis of conventional Western medicine treatment, which is under the guidance of Syndrome differentiation and treatment. To avoid irregularities in medication administration, each patient in the combined treatment group was prescribed Chinese herbal medicines by the doctor in charge, during which the prescription was adjusted according to the changes in the patient's symptoms. This also reflects the individualized and dynamic evolution of TCM in real-world studies. To control the influence of confounding factors on the results due to the non-randomized trial design, we applied PSM. In addition, physical activity, the Gensini score, the SAQ score, and other observations were included, as these indicators can objectively reflect the disease status of patients with SAP of CHD and are closely related to the prognosis of cardiovascular disease –35.Nonetheless, this study has limitations. The follow-up period of this trial was short and only reflected the clinical effectiveness and safety of drug administration in subjects over a 1-year period. In addition, the single-center design of the trial allowed for a limited sample size and insufficient external validity of the model. In addition, the problem of heterogeneity in the combined drug group cannot be solved yet, which is one of the objective limitations of TCM in real-world studies. In the future, a multicenter, large-sample real-world research will be required to enhance the external validity of our model. Meanwhile, the follow-up period should be appropriately prolonged to further validate the effectiveness of integrated Chinese and Western medicine on the long-term prognosis of patients with SAP of CHD. Additionally, we will record each patient's prescription for medication in detail in future studies and consider setting up subgroup analyses based on the specific use of herbal medicines by patients to improve the credibility of the article.5.We investigated the effect of real-world combined Chinese and Western medical therapy vs. conventional therapy on patient outcome events in patients with stable angina pectoris. We also developed multifactor logistic regression tests to explore the risk factors for the occurrence of stable angina pectoris in patients. Ultimately, we conclude that treatment modality is an independent factor influencing the occurrence of CVEs in patients with stable angina pectoris of coronary artery disease. Integrated treatment based on Chinese and Western medicine might improve the prognosis and reduce the risk of CVEs in this disease population.